Detection of Pathogenesis-Related (PR) Genes in Susceptible Tomato Plants Infected by Meloidogyne spp.

Meloidogyne species, as infective second-stage juveniles (J2s) larvae, are parasites able to attack host of relevant agronomic interest such as tomato plants. The identification of gene expression markers, useful to investigate the levels of root-knot nematode infection in the roots, is a fundamental tool in plant-pathogen interaction. The laboratory methods for analyzing the differential expression of pathogenesis-related (PR) genes constitute powerful tools for detecting the induced systemic acquired resistance defense response to M. incognita in infected plants and can be extended to all pathogen infection markers to obtain an early and sustainable control.

Distribution of Small RNAs Along Transposable Elements in Vitis vinifera During Somatic Embryogenesis.

Metaviridae is a family of reverse-transcribing viruses, closely related to retroviruses; they exist within their host's DNA as transposable elements. Transposable element study requires the use of specialized tools, in part because of their repetitive nature. By combining data from transcript RNA-Seq, small RNA-Seq, and parallel analysis of RNA ends-Seq from grapevine somatic embryos, we set up a bioinformatics flowchart that could be able to assemble and identify transposable elements.

Resistance to Plant Parasites in Tomato Is Induced by Soil Enrichment with Specific Bacterial and Fungal Rhizosphere Microbiome.

Commercial formulations of beneficial microbes have been used to enrich the rhizosphere microbiome of tomato plants grown in pots located in a glasshouse. These plants have been subjected to attacks by soil-borne parasites, such as root-knot nematodes (RKNs), and herbivores, such as the miner insect Tuta absoluta. The development of both parasites and the symptoms of their parasitism were restricted in these plants with respect to plants left untreated. A mixture, named in the text as Myco, containing plant growth-promoting rhizobacteria (PGPR), opportunistic biocontrol fungi (BCF), and arbuscular mycorrhizal fungi (AMF) was more effective in limiting pest damage than a formulation containing the sole AMF (Ozor). Therefore, Myco-treated plants inoculated with RKNs were taken as a model for further studies. The PGPR contained in Myco were not able to reduce nematode infection; rather, they worsened symptoms in plants compared with those observed in untreated plants. Therefore, it was argued that both BCF and AMF were the microorganisms that colonized roots and stimulated the plant immune system against RKNs. Beneficial fungi colonized the roots by lowering the activities of the defense supporting enzymes endochitinases and ß-1,3-glucanase. However, as early as three days after nematode inoculation, these enzyme activities and the expression of the encoding pathogenesis-related genes (PR-2, PR-3) were found to be enhanced in roots with respect to non-inoculated plants, thus indicating that plants had been primed against RKNs. The addition of paclobutrazol, which reduces salicylic acid (SA) levels in cells, and diphenyliodonium chloride, which inhibits superoxide generation, completely abolished the repressive effect of Myco on nematode infection. Inhibitors of copper enzymes and the alternative cyanide-resistant respiration did not significantly alter resistance induction by Myco. When Myco-treated plants were subjected to moderate water stress and inoculated with nematodes, they retained numbers of developed individuals in the roots similar to those present in regularly watered plants, in contrast to what occurred in roots of untreated stressed plants that hosted very few individuals because of poor nutrient availability.

Exogenous and endogenous dsRNAs perceived by plant Dicer-like 4 protein in the RNAi-depleted cellular context.

BACKGROUND: In plants, RNase III Dicer-like proteins (DCLs) act as sensors of dsRNAs and process them into short 21- to 24-nucleotide (nt) (s)RNAs. Plant DCL4 is involved in the biogenesis of either functional endogenous or exogenous (i.e. viral) short interfering (si)RNAs, thus playing crucial antiviral roles. METHODS: In this study we expressed plant DCL4 in Saccharomyces cerevisiae, an RNAi-depleted organism, in which we could highlight the role of dicing as neither Argonautes nor RNA-dependent RNA polymerase is present. We have therefore tested the DCL4 functionality in processing exogenous dsRNA-like substrates, such as a replicase-assisted viral replicon defective-interfering RNA and RNA hairpin substrates, or endogenous antisense transcripts. RESULTS: DCL4 was shown to be functional in processing dsRNA-like molecules in vitro and in vivo into 21- and 22-nt sRNAs. Conversely, DCL4 did not efficiently process a replicase-assisted viral replicon in vivo, providing evidence that viral RNAs are not accessible to DCL4 in membranes associated in active replication. Worthy of note, in yeast cells expressing DCL4, 21- and 22-nt sRNAs are associated with endogenous loci. CONCLUSIONS: We provide new keys to interpret what was studied so far on antiviral DCL4 in the host system. The results all together confirm the role of sense/antisense RNA-based regulation of gene expression, expanding the sense/antisense atlas of S. cerevisiae. The results described herein show that S. cerevisiae can provide insights into the functionality of plant dicers and extend the S. cerevisiae tool to new biotechnological applications.

Inhibition of ROS-Scavenging Enzyme System Is a Key Event in Tomato Genetic Resistance against Root-Knot Nematodes.

Genetic resistance in plants against incompatible pests is expressed by the activation of an immune system; however, the molecular mechanisms of pest recognition and expression of immunity, although long the object of investigation, are far from being fully understood. The immune response triggered by the infection of soil-borne parasites, such as root-knot nematodes (RKNs), to incompatible resistant tomato plants was studied and compared to the compatible response that occurred when RKNs attacked susceptible plants. In compatible interactions, the invading nematode juveniles were allowed to fully develop and reproduce, whilst that was impeded in incompatible interactions. In crude root extracts, a first assay of reactive oxygen species (ROS)-scavenging enzymatic activity was carried out at the earliest stages of tomato-RKN incompatible interaction. Membrane-bound and soluble CAT, which is the most active enzyme in hydrogen peroxide (H2O2) scavenging, was found to be specifically inhibited in roots of inoculated resistant plants until 5 days after inoculation, with respect to uninoculated plants. The expression of genes encoding for antioxidant enzymes, such as CAT and glutathione peroxidase (GPX), was not always inhibited in roots of nematode-infected resistant tomato. Therefore, the biochemical mechanisms of CAT inhibition were further investigated. Two CAT isozymes were characterized by size exclusion HPLC as a tetrameric form with a molecular weight of 220,000 dalton and its subunits (55,000 dalton). Fractions containing such isozymes were tested by their sensitivity to both salicylic acid (SA) and H2O2. It was evidenced that elevated concentrations of both chemicals led to a partial inactivation of CAT. Elevated concentrations of H2O2 in incompatible interactions have been suggested to be produced by membrane-bound superoxide anion generating, SOD, and isoperoxidase-enhanced activities. Such partial inactivation of CAT has been depicted as one of the earliest key metabolic events, which is specifically associated with tomato immunity to RKNs. Enhanced ROS production and the inhibition of ROS-scavenging systems have been considered to trigger all the metabolic events leading to cell death and tissue necrosis developed around the head of the invading juveniles by which this special type of plant resistance is exerted.

Tomato Root Colonization by Exogenously Inoculated Arbuscular Mycorrhizal Fungi Induces Resistance against Root-Knot Nematodes in a Dose-Dependent Manner.

Arbuscular mycorrhizal fungi (AMF) are generally recognized to induce plant growth and prime plants against soil-borne parasites, such as plant parasitic nematodes. However, the effectiveness of commercial formulates containing AMF has been questioned. Increasing amounts per plant of one commercial AMF-containing formulate, reported in the text as Myco, were used to detect the effects on growth of tomato plants and the resistance induced against root-knot nematodes (RKNs) The doses used per plant (0.5, 1.0, 2.0 g, reported as Myco1, Myco2, Myco3, respectively) were soil-drenched to growing potted plants; the effects of such treatments were analyzed both in plants not inoculated or inoculated by Meloidogyne incognita juveniles. Consistent increases in plant weight were apparent as soon as 7 days only after Myco2 treatments. Moreover, only treatments with Myco2 induced a consistent repression of the nematode infection observed in untreated plants. Conversely, treatments with Myco1 and Myco3 did not produce such an early growth improvement; some plant weight increase was observable only at 28 dpt. Accordingly, such Myco doses did not restrict the level of infestation observed in untreated plants. Control of infection was dependent on the dose of Myco provided to plants five days before nematode inoculation. About one month after all Myco treatments, several areas of roots were found to be colonized by AMF, although in Myco2-treated plants, three genes involved in the AMF colonization process (SlCCaMK, SlLYK9, and SlLYK13) were found to be over-expressed already at 7 dpt; over-expression was generally less consistent at 14 and 21 dpt. The expressions of two key genes of plant defense, the hypersensitive cell death inducer PR4b gene and the glutathione peroxidase-encoding GPX gene, were monitored in roots of Myco2-treated plants 3 and 7 days after nematode inoculation. PR4b was over-expressed and GPX was silenced in treated plants with respect to untreated plants. The repressive effect of Myco2 treatment against RKN infection was completely abolished when Myco2 suspensions were autoclaved to sterilization or treated with the potent anti-fungal agent amphotericin B, thus indicating that the biological control agents contained in the commercial formulate were living fungi.

Identification of Known and Novel Arundo donax L. MicroRNAs and Their Targets Using High-Throughput Sequencing and Degradome Analysis.

MicroRNAs (miRNAs) are a class of non-coding molecules involved in the regulation of a variety of biological processes. They have been identified and characterized in several plant species, but only limited data are available for Arundo donax L., one of the most promising bioenergy crops. Here we identified, for the first time, A. donax conserved and novel miRNAs together with their targets, through a combined analysis of high-throughput sequencing of small RNAs, transcriptome and degradome data. A total of 134 conserved miRNAs, belonging to 45 families, and 27 novel miRNA candidates were identified, along with the corresponding primary and precursor miRNA sequences. A total of 96 targets, 69 for known miRNAs and 27 for novel miRNA candidates, were also identified by degradome analysis and selected slice sites were validated by 5'-RACE. The identified set of conserved and novel candidate miRNAs, together with their targets, extends our knowledge about miRNAs in monocots and pave the way to further investigations on miRNAs-mediated regulatory processes in A. donax, Poaceae and other bioenergy crops.

Composted Municipal Green Waste Infused with Biocontrol Agents to Control Plant Parasitic Nematodes-A Review.

The last few years have witnessed the emergence of alternative measures to control plant parasitic nematodes (PPNs). We briefly reviewed the potential of compost and the direct or indirect roles of soil-dwelling organisms against PPNs. We compiled and assessed the most intensively researched factors of suppressivity. Municipal green waste (MGW) was identified and profiled. We found that compost, with or without beneficial microorganisms as biocontrol agents (BCAs) against PPNs, were shown to have mechanisms for the control of plant parasitic nematodes. Compost supports a diverse microbiome, introduces and enhances populations of antagonistic microorganisms, releases nematicidal compounds, increases the tolerance and resistance of plants, and encourages the establishment of a "soil environment" that is unsuitable for PPNs. Our compilation of recent papers reveals that while the scope of research on compost and BCAs is extensive, the role of MGW-based compost (MGWC) in the control of PPNs has been given less attention. We conclude that the most environmentally friendly and long-term, sustainable form of PPN control is to encourage and enhance the soil microbiome. MGW is a valuable resource material produced in significant amounts worldwide. More studies are suggested on the use of MGWC, because it has a considerable potential to create and maintain soil suppressivity against PPNs. To expand knowledge, future research directions shall include trials investigating MGWC, inoculated with BCAs.

Regulation of plant antiviral defense genes via host RNA-silencing mechanisms.

BACKGROUND: Plants in nature or crops in the field interact with a multitude of beneficial or parasitic organisms, including bacteria, fungi and viruses. Viruses are highly specialized to infect a limited range of host plants, leading in extreme cases to the full invasion of the host and a diseased phenotype. Resistance to viruses can be mediated by various passive or active mechanisms, including the RNA-silencing machinery and the innate immune system. MAIN TEXT: RNA-silencing mechanisms may inhibit viral replication, while viral components can elicit the innate immune system. Viruses that successfully enter the plant cell can elicit pattern-triggered immunity (PTI), albeit by yet unknown mechanisms. As a counter defense, viruses suppress PTI. Furthermore, viral Avirulence proteins (Avr) may be detected by intracellular immune receptors (Resistance proteins) to elicit effector-triggered immunity (ETI). ETI often culminates in a localized programmed cell death reaction, the hypersensitive response (HR), and is accompanied by a potent systemic defense response. In a dichotomous view, RNA silencing and innate immunity are seen as two separate mechanisms of resistance. Here, we review the intricate connections and similarities between these two regulatory systems, which are collectively required to ensure plant fitness and resilience. CONCLUSIONS: The detailed understanding of immune regulation at the transcriptional level provides novel opportunities for enhancing plant resistance to viruses by RNA-based technologies. However, extensive use of RNA technologies requires a thorough understanding of the molecular mechanisms of RNA gene regulation. We describe the main examples of host RNA-mediated regulation of virus resistance.

Endogenous activated small interfering RNAs in virus-infected Brassicaceae crops show a common host gene-silencing pattern affecting photosynthesis and stress response.

Viral infections are accompanied by a massive production of small interfering RNAs (siRNAs) of plant origin, such as virus-activated (va)siRNAs, which drive the widespread silencing of host gene expression, and whose effects in plant pathogen interactions remain unknown. By combining phenotyping and molecular analyses, we characterized vasiRNAs that are associated with typical mosaic symptoms of cauliflower mosaic virus infection in two crops, turnip (Brassica rapa) and oilseed rape (Brassica napus), and the reference plant Arabidopsis thaliana. We identified 15 loci in the three infected plant species, whose transcripts originate vasiRNAs. These loci appear to be generally affected by virus infections in Brassicaceae and encode factors that are centrally involved in photosynthesis and stress response, such as Rubisco activase (RCA), senescence-associated protein, heat shock protein HSP70, light harvesting complex, and membrane-related protein CP5. During infection, the expression of these factors is significantly downregulated, suggesting that their silencing is a central component of the plant's response to virus infections. Further findings indicate an important role for 22 nt long vasiRNAs in the plant's endogenous RNA silencing response. Our study considerably enhances knowledge about the new class of vasiRNAs that are triggered in virus-infected plants and will help to advance strategies for the engineering of gene clusters involved in the development of crop diseases.

Epigenetic and Metabolic Changes in Root-Knot Nematode-Plant Interactions.

Two wild-type field populations of root-knot nematodes (Mi-Vfield, Mj-TunC2field), and two isolates selected for virulence in laboratory on resistant tomato cultivars (SM2V, SM11C2), were used to induce a resistance reaction in tomato to the soil-borne parasites. Epigenetic and metabolic mechanisms of resistance were detected and compared with those occurring in partially or fully successful infections. The activated epigenetic mechanisms in plant resistance, as opposed to those activated in infected plants, were detected by analyzing the methylated status of total DNA, by ELISA methods, and the expression level of key genes involved in the methylation pathway, by qRT-PCR. DNA hypo-methylation and down-regulation of two methyl-transferase genes (CMT2, DRM5), characterized the only true resistant reaction obtained by inoculating the Mi-1.2-carrying resistant tomato cv Rossol with the avirulent field population Mi-Vfield. On the contrary, in the roots into which nematodes were allowed to develop and reproduce, total DNA was generally found to be hyper-methylated and methyl-transferase genes up-loaded. DNA hypo-methylation was considered to be the upstream mechanism that triggers the general gene over-expression observed in plant resistance. Gene silencing induced by nematodes may be obtained through DNA hyper-methylation and methyl-transferase gene activation. Plant resistance is also characterized by an inhibition of the anti-oxidant enzyme system and activation of the defense enzyme chitinase, as opposed to the activation of such a system and inhibition of the defense enzyme glucanase in roots infested by nematodes.

Plant miRNA Cross-Kingdom Transfer Targeting Parasitic and Mutualistic Organisms as a Tool to Advance Modern Agriculture.

MicroRNAs (miRNAs), defined as small non-coding RNA molecules, are fine regulators of gene expression. In plants, miRNAs are well-known for regulating processes spanning from cell development to biotic and abiotic stress responses. Recently, miRNAs have been investigated for their potential transfer to distantly related organisms where they may exert regulatory functions in a cross-kingdom fashion. Cross-kingdom miRNA transfer has been observed in host-pathogen relations as well as symbiotic or mutualistic relations. All these can have important implications as plant miRNAs can be exploited to inhibit pathogen development or aid mutualistic relations. Similarly, miRNAs from eukaryotic organisms can be transferred to plants, thus suppressing host immunity. This two-way lane could have a significant impact on understanding inter-species relations and, more importantly, could leverage miRNA-based technologies for agricultural practices. Additionally, artificial miRNAs (amiRNAs) produced by engineered plants can be transferred to plant-feeding organisms in order to specifically regulate their cross-kingdom target genes. This minireview provides a brief overview of cross-kingdom plant miRNA transfer, focusing on parasitic and mutualistic relations that can have an impact on agricultural practices and discusses some opportunities related to miRNA-based technologies. Although promising, miRNA cross-kingdom transfer remains a debated argument. Several mechanistic aspects, such as the availability, transfer, and uptake of miRNAs, as well as their potential to alter gene expression in a cross-kingdom manner, remain to be addressed.

Viral and subviral derived small RNAs as pathogenic determinants in plants and insects.

The phenotypic manifestations of disease induced by viruses and subviral infectious entities are the result of complex molecular interactions between host and viral factors. The viral determinants of the diseased phenotype have traditionally been sought at the level of structural or non-structural proteins. However, the discovery of RNA silencing mechanisms has led to speculations that determinants of the diseased phenotype are caused by viral nucleic acid sequences in addition to proteins. RNA silencing is a gene regulation mechanism conserved within eukaryotic kingdoms (with the exception of some yeast species), and in plants and insects it also functions as an antiviral mechanism. Non-coding RNAs of viral origin, ranging in size from 21 to 24 nucleotides (viral small interfering RNAs, vsiRNAs) accumulate in virus-infected tissues and organs, in some cases to comparable levels as the entire complement of host-encoded small interfering RNAs. Upon incorporation into RNA-induced silencing complexes, vsiRNAs can mediate cleavage or induce translational inhibition of nucleic acid targets in a sequence-specific manner. This review focuses on recent findings that suggest an increased complexity of small RNA-based interactions between virus and host. We mainly address plant viruses, but where applicable discuss insect viruses as well. Prominence is given to studies that have indisputably demonstrated that vsiRNAs determine diseased phenotype by either carrying sequence determinants or, indirectly, by altering host-gene regulatory pathways. Results from these studies suggest biotechnological applications, which are also discussed.

Bio-control agents activate plant immune response and prime susceptible tomato against root-knot nematodes.

Beneficial microorganisms are generally known to activate plant defense against biotic challenges. However, the molecular mechanisms by which activated plants react more rapidly and actively to pests remain still largely unclear. Tomato plants pre-treated with a mixture of beneficial bio-control agents (BCAs), as soil-drenches, were less sensitive to infection of the root-knot nematode (RKN) Meloidogyne incognita. To unravel the molecular mechanisms of this induced resistance against RKNs, we used qRT-PCR to monitor the expression, in tomato roots and leaves, of 6 key defense genes. Gene transcripts were detected until the 12th day after BCA treatment(3, 7, 8, 12 dpt) and3 and 7 days after nematode inoculation of pre-treated plants. Early after BCA treatment, the salicylic acid (SA)-dependent pathogenesis related gene (PR-gene), PR-1b, marker of the systemic acquired resistance (SAR), was systemically over-expressed. Another PR-gene, PR-5, was over-expressed at later stages of BCA-plant interaction, and only in roots. Activation of defense against RKNs was attested by the early up-regulation of 4 genes (PR-1, PR-3, PR-5, ACO) in pre-treated plants after inoculation. Conversely, the expression of the JA/ET-dependent gene JERF3 did not increase after nematode inoculation in primed plants. A catalase gene (CAT)was highly over-expressed by nematode infection, however, this over-expression was annulled at the earliest stages or limited at the later stages of infection toBCA-treated roots. Enzyme activities, such as glucanase and endochitinase, were enhanced in roots of pre-treated inoculated plants with respect to plants left not inoculated as a control. These findings indicate that BCA interaction with roots primes plants against RKNs. BCA-mediated immunity seems to rely on SA-mediated SAR and to be associated with both the activation of chitinase and glucanase enzyme activities and the inhibition of the plant antioxidant enzyme system. Immunity is triggered at the penetration and movements inside the roots of the invading nematode juveniles but probably acts at the feeding site building stage of nematode infection.

Viruses and Phytoparasitic Nematodes of Cicer arietinum L.: Biotechnological Approaches in Interaction Studies and for Sustainable Control.

Cicer arietinum L. (chickpea) is the world's fourth most widely grown pulse. Chickpea seeds are a primary source of dietary protein for humans, and chickpea cultivation contributes to biological nitrogen fixation in the soil, given its symbiotic relationship with rhizobia. Therefore, chickpea cultivation plays a pivotal role in innovative sustainable models of agro-ecosystems inserted in crop rotation in arid and semi-arid environments for soil improvement and the reduction of chemical inputs. Indeed, the arid and semi-arid tropical zones of Africa and Asia have been primary areas of cultivation and diversification. Yet, nowadays, chickpea is gaining prominence in Canada, Australia, and South America where it constitutes a main ingredient in vegetarian and vegan diets. Viruses and plant parasitic nematodes (PPNs) have been considered to be of minor and local impact in primary areas of cultivation. However, the introduction of chickpea in new environments exposes the crop to these biotic stresses, compromising its yields. The adoption of high-throughput genomic technologies, including genome and transcriptome sequencing projects by the chickpea research community, has provided major insights into genome evolution as well as genomic architecture and domestication. This review summarizes the major viruses and PPNs that affect chickpea cultivation worldwide. We also present an overview of the current state of chickpea genomics. Accordingly, we explore the opportunities that genomics, post-genomics and novel editing biotechnologies are offering in order to understand chickpea diseases and stress tolerance and to design innovative control strategies.

ROS and Cell Death in Tomato Roots Infected by Meloidogyne Incognita.

Phytoparasitic nematodes are plant pests causing serious problems to a broad range of hosts, and Meloidogyne species are widely recognized as the most damaging among the root knot nematode groups. During the incompatible interaction between avirulent pathogens and resistant tomato cultivars, juvenile nematode invasions provoke a defense cascade, culminating in hypersensitive responses. Methods to detect the key molecules involved in oxidative metabolism of the infected tomato roots are described here.

The Seed Repair Response during Germination: Disclosing Correlations between DNA Repair, Antioxidant Response, and Chromatin Remodeling in Medicago truncatula.

This work provides novel insights into the effects caused by the histone deacetylase inhibitor trichostatin A (TSA) during Medicago truncatula seed germination, with emphasis on the seed repair response. Seeds treated with H2O and TSA (10 and 20 µM) were collected during imbibition (8 h) and at the radicle protrusion phase. Biometric data showed delayed germination and impaired seedling growth in TSA-treated samples. Comet assay, performed on radicles at the protrusion phase and 4-days old M. truncatula seedlings, revealed accumulation of DNA strand breaks upon exposure to TSA. Activation of DNA repair toward TSA-mediated genotoxic damage was evidenced by the up-regulation of MtOGG1(8-OXOGUANINE GLYCOSYLASE/LYASE) gene involved in the removal of oxidative DNA lesions, MtLIGIV(LIGASE IV) gene, a key determinant of seed quality, required for the rejoining of DNA double strand breaks and TDP(TYROSYL-DNA PHOSPHODIESTERASE) genes encoding the multipurpose DNA repair enzymes tyrosyl-DNA phosphodiesterases. Since radical scavenging can prevent DNA damage, the specific antioxidant activity (SAA) was measured by DPPH (1,1-diphenyl-2-picrylhydrazyl) and Folin-Ciocalteu reagent assays. Fluctuations of SAA were observed in TSA-treated seeds/seedlings concomitant with the up-regulation of antioxidant genes MtSOD(SUPEROXIDE DISMUTASE, MtAPX(ASCORBATE PEROXIDASE) and MtMT2(TYPE 2 METALLOTHIONEIN). Chromatin remodeling, required to facilitate the access of DNA repair enzymes at the damaged sites, is also part of the multifaceted seed repair response. To address this aspect, still poorly explored in plants, the MtTRRAP(TRANSFORMATION/TRANSACTIVATION DOMAIN-ASSOCIATED PROTEIN) gene was analyzed. TRRAP is a transcriptional adaptor, so far characterized only in human cells where it is needed for the recruitment of histone acetyltransferase complexes to chromatin during DNA repair. The MtTRRAP gene and the predicted interacting partners MtHAM2 (HISTONE ACETYLTRANSFERASE OF THE MYST FAMILY) and MtADA2A (TRANSCRIPTIONAL ADAPTOR) showed tissue- and dose-dependent fluctuations in transcript levels. PCA (Principal Component Analysis) and correlation analyses suggest for a new putative link between DNA repair and chromatin remodeling that involves MtOGG1 and MtTRRAP genes, in the context of seed germination. Interesting correlations also connect DNA repair and chromatin remodeling with antioxidant players and proliferation markers.

Induction of SA-signaling pathway and ethylene biosynthesis in Trichoderma harzianum-treated tomato plants after infection of the root-knot nematode Meloidogyne incognita.

KEY MESSAGE: Salicylic acid-signaling pathway and ethylene biosynthesis were induced in tomato treated with Trichoderma harzianum when infected by root-knot nematodes and limited the infection by activation of SAR and ethylene production. Soil pre-treatment with Trichoderma harzianum (Th) strains ITEM 908 (T908) and T908-5 decreased susceptibility of tomato to Meloidogyne incognita, as assessed by restriction in nematode reproduction and development. The effect of T. harzianum treatments on plant defense was detected by monitoring the expression of the genes PR-1/PR-5 and JERF3/ACO, markers of the SA- and JA/ET-dependent signaling pathways, respectively. The compatible nematode-plant interaction in absence of fungi caused a marked suppression of PR-1, PR-5, and ACO gene expressions, either locally or systemically, whilst expression of JERF3 gene resulted unaffected. Conversely, when plants were pre-treated with Th-strains, over-expression of PR-1, PR-5, and ACO genes was observed in roots 5 days after nematode inoculation. JERF3 gene expression did not change in Th-colonized plants challenged with nematodes. In the absence of nematodes, Trichoderma-root interaction was characterized by the inhibition of both SA-dependent signaling pathway and ET biosynthesis, and, in the case of PR-1 and ACO genes, this inhibition was systemic. JERF3 gene expression was systemically restricted only at the very early stages of plant-fungi interaction. Data presented indicate that Th-colonization primed roots for Systemic Acquired Resistance (SAR) against root-knot nematodes and reacted to nematode infection more efficiently than untreated plants. Such a response probably involves also activation of ET production, through an augmented transcription of the ACO gene, which encodes for the enzyme catalyzing the last step of ET biosynthesis. JA signaling and Induced Systemic Resistance (ISR) do not seem to be involved in the biocontrol action of the tested Th-strains against RKNs.