Spot Disease Of Tomato Research Articles

The Control of Pseudomonas syringae pv. tomato, Causing Bacterial Spot Disease in Tomato, by Different Plant Essential Oils

Seed-borne Pseudomonas syringae pv. tomato causes bacterial spot disease in tomatoes and causes serious economic losses by affecting yield and quality in tomato cultivation. In this study, the effect of essential oils obtained by distillation from Mentha longifolia, Thymus migricus, Salvia hydrangea and Artemisia absinthium and Achillea arabica plants on the control of P. syringae pv. tomato was investigated. The antibacterial activities of plant essential oils against P. syringae pv. tomato were tested by the in vitro agar disk diffusion method. Additionally, minimum inhibitory concentrations at which plant essential oils inhibit the growth of Pst 76 were determined. It was determined that all oils created an inhibition zone between 4.0-30.0 mm against the disease agent and the highest activity was obtained from application with the oil of the T. migricus plant. It was determined that the oils obtained from T. migricus and M. longifolia plants had higher antimicrobial activity than methylmycin in the control group. In line with the results obtained, disease control with essential oils has been seen as an alternative method that can be included in integrated control programs.

Photosensitizer to the rescue: in planta and field application of photodynamic inactivation against plant pathogenic bacteria.

Control of plant pathogens using chemical and synthetic pesticides raises a major safety concern for humans and the environment. Despite the ongoing exploration of sustainable alternative methods, management practices for pathogens, especially bacteria, have remained almost unchanged over decades, whereby long-term uses of copper and antibiotics has led to widespread bacterial resistance in the field. Antimicrobial photodynamic inactivation (aPDI) of bacteria is emerging as an alternative strategy to combat resistant plant pathogens. aPDI utilizes light-sensitive molecules (photosensitizers) that upon illumination produce reactive oxygen species able to kill pathogens. Here we explore the potential of an anionic semisynthetic water-soluble derivative of chlorophyl (Sodium Magnesium Chlorophyllin: Mg-chl), as an antibacterial agent in planta, by simulating processes naturally occurring in the field. Mg-chl in combination with Na2EDTA (cell wall permeabilizing agent) was able to effectively inhibit Pseudomonas syringae pv. tomato DC3000 in vitro and in planta in both tomato and N. benthamiana. Notably, Mg-chl in combination with Na2EDTA and the common surfactant Morwet D-400 significantly reduced Xanthomonas hortorum pv. gardneri and Xanthomonas fragarie, respectively, in a commercial greenhouse trial against bacterial spot disease in tomato and in field experiments against angular leaf spot disease in strawberries.

Survey and Molecular Identification of Xanthomonas euvesicatoria Causing Bacterial Spot of Tomato in Blue Nile State in Sudan

This study aimed to survey, characterize, and identify the causal pathogen of tomato bacterial spots (TBS). The survey evaluated the disease incidence and severity on tomato cultivated in Blue Nile State during the autumn season 2015. The surveyed sites were in Alrosseris and Aldmazein localities. Four farms were selected from each locality. Symptomatic leaves were collected and sent to plant pathology laboratory, Department of Crop protection, Faculty of Agriculture, University of Khartoum for detection of the pathogen. Pathogenicity of the isolated bacteria was confirmed by relevant test. The pathogen was isolated and characterized morphologically, biochemically and at molecular level. The survey revealed that there was no significant difference in disease incidence within each locality, while there were significant differences (P ≤ 0.05) among locations within both localities in disease severity. The disease incidence was remarkably high in Alrosseris and Aldamazein which recorded 49.27 and 52.17 %, respectively, while the disease severity was 28.28 and 30.32%, respectively. The results showed that the isolates were rod shaped, obligate aerobic, Gram negative, catalase positive, oxidase negative, and were able to hydrolyse starch and gelatine, yellow colonies on nutrient agar and Yeast Dextrose Calcium carbonate (YDC) media. In the pathogenicity test, the bacterium known as Xanthomonas vesicatoria was confirmed to be as Xanthomonas euvesicatoria by PCR and sequence analysis of the 16S RNA gene (Gen Bank Accession No's MH047248, MH051263 and MG686236).These results also indicate that the tomato plants grown in Blue Nile State were highly infected by the bacterial leaf spots. Further studies should be considered to determine potential yield loss caused by the pathogen, as well as control strategies to limit the spread of this pathogen. To our knowledge, this is the first report of Xanthomonas euvesicatoria causing bacterial spot disease in tomato in Sudan

Stemphylium lycopersici Nep1-like Protein (NLP) Is a Key Virulence Factor in Tomato Gray Leaf Spot Disease.

The fungus Stemphylium lycopersici (S. lycopersici) is an economically important plant pathogen that causes grey leaf spot disease in tomato. However, functional genomic studies in S. lycopersici are lacking, and the factors influencing its pathogenicity remain largely unknown. Here, we present the first example of genetic transformation and targeted gene replacement in S. lycopersici. We functionally analyzed the NLP gene, which encodes a necrosis- and ethylene-inducing peptide 1 (Nep1)-like protein (NLP). We found that targeted disruption of the NLP gene in S. lycopersici significantly compromised its virulence on tomato. Moreover, our data suggest that NLP affects S. lycopersici conidiospore production and weakly affects its adaptation to osmotic and oxidative stress. Interestingly, we found that NLP suppressed the production of reactive oxygen species (ROS) in tomato leaves during S. lycopersici infection. Further, expressing the fungal NLP in tomato resulted in constitutive transcription of immune-responsive genes and inhibited plant growth. Through gene manipulation, we demonstrated the function of NLP in S. lycopersici virulence and development. Our work provides a paradigm for functional genomics studies in a non-model fungal pathogen system.

First Report of Xanthomonas euvesicatoria Causing Bacterial Spot Disease in Tomato in Blue Nile State, Sudan

Bacterial spot caused by Xanthomonas vesicatoria is considered to be a major constraint of tomato production all over the world in particular warmer and humid parts.A disease disorder was reported in autumn season 2015within the Blue Nile State in Sudanaffecting all aerial parts of tomato plant. Symptomatic tomato leaves were collected and sent toPlant Pathology Laboratory, Faculty of Agriculture, University of Khartoum. An investigation on whether a phytopathogen was associated with symptoms of bacterial spots damage on tomato was conducted.The pathogen wasisolated and characterized morphologically, biochemically and at molecular level. The bacterium wasidentified as Xanthomonaseuvesicatoria and confirmed by PCR and sequence analysis of the 16S RNA gene (GenBankAccession Nos MH047248, MH051263 and MG686236).The disease incidence was remarkably high in ALrosserisand Aldamazein which recorded 49.27 and 52.17%, respectively, while the disease severity was 28.28 and 30.32%, respectively. To our knowledge, this is the first report of Xanthomonaseuvesicatoria causing bacterial spot disease in tomato in Sudan.

Evaluation of homeopathic drugs for the management of Alternaria solani causing leaf spot disease in tomato under controlled conditions

Man is dependent on plants for almost every need and requirement. Hence, destruction of crop plants due to infection by fungal pathogens has always been an area of prime concern. The use of synthetic fungicides for disease management is harmful for human and animal health as well as soil. They enter the food chain and cause several deleterious effects on biosphere, contributing to significant declines in populations of beneficial soil organisms, soil acidification and compaction, thatch accumulation, and diminished resistance to diseases. Natural plant products are important sources of new agrochemicals for the control of plant diseases. Therefore, the In vitro evaluation of Homeopathic drugs against Alternaria solani was carried out at M.P.K.V, Rahuri. Total 20 different Homeopathic drugs were evaluated at 10%, 15% and 20% concentration by Poison food technique against these pathogens. Among the 20 different homeopathic drugs, Sulphur, Pulsatila nigricans, Podophyllum pellatum, Cina, Lycopodium clavatum, Nux vomica, Dulcamara, Colocynthis, Aconitum napellun, Natrum muriatum, Rhux toxicodendron, Arnica montana, Hepar sulphur, Arsenicum album were showed 100% inhibition over the growth of Alternaria solani were effective at 20% concentration. The drugs like Apis melifera, Cinchona officinalis, Sepia and Calcaria carbonica showed 87.78, 80.00, 64.44 and 57.78 % inhibition over Alternaria solani respectively.

A sulfated polysaccharide κ-carrageenan induced antioxidant defense and proteomic changes in chloroplast against leaf spot disease of tomato

Ecofriendly approaches to control plant diseases using biopolymers are gaining momentum due to their non-toxic nature to the environment. The present study elucidates the potency of a sulfated polysaccharide, κ-carrageenan, obtained from the red seaweed Kappaphycus alvarezii, as an inducer of antioxidant defense and also chloroplast proteome against leaf spot disease in tomato. к-carrageenan (0.3%) treatment was very effective in reducing disease scores in tomato plants. Fluorescent microscopy of tomato leaves stained with Uvitex 2B (a chitin specific stain) revealed that foliar application of κ-carrageenan (0.3%) reduced colonization of the pathogen Septoria lycopersici. In addition, κ-carrageenan pretreatment followed by pathogen infection recorded a higher accumulation of calcium oxalate crystals. The treatment of κ-carrageenan resulted in the induction of H2O2 and superoxide anion compared to control or S. lycopersici-infected plants. The tomato leaves treated with κ-carrageenan showed higher activity of peroxidase enzyme. Furthermore, chloroplast proteome analysis by 2D-PAGE of control, S. lycopersici-infected, κ-carrageenan-treated, and к-carrageenan treatment followed by S. lycopersici-infected leaves of tomato showed differential expression of 31 proteins, of which 11 proteins were upregulated including acid phosphatase 1-like, C- terminal processing peptidase, GATA transcription factor 5, Calvin cycle protein CP12-1, Photosystem I reaction center subunit PSAK, and subunit of cytochrome B6-F complex iron-sulfur centers. These results indicated that κ-carrageenan could be a promising inducer of antioxidant defense and modulator of chloroplast proteome in tomato plants.

Characterization of three novel genetic loci encoding bacteriocins associated with Xanthomonasperforans.

Bacterial spot is a destructive disease of tomato in Florida that prior to the early 1990s was caused by Xanthomonas euvesicatoria. X. perforans was first identified in Florida in 1991 and by 2006 was the only xanthomonad associated with bacterial spot disease in tomato. The ability of an X. perforans strain to outcompete X. euvesicatoria both in vitro and in vivo was at least in part associated with the production of three bacteriocins designated Bcn-A, Bcn-B, and Bcn-C. The objective of this study was to characterize the genetic determinants of these bacteriocins. Bcn-A activity was confined to one locus consisting of five ORFs of which three (ORFA, ORF2 and ORF4) were required for bacteriocin activity. The fifth ORF is predicted to encode an immunity protein to Bcn-A based on in vitro and in vivo assays. The first ORF encodes Bcn-A, a 1,398 amino acid protein, which bioinformatic analysis predicts to be a member of the RHS family of toxins. Based on results of homology modeling, we hypothesize that the amino terminus of Bcn-A interacts with a protein in the outer membrane of X. euvesicatoria. The carboxy terminus of the protein may interact with an as yet unknown protein(s) and puncture the X. euvesicatoria membrane, thereby delivering the accessory proteins into the target and causing cell death. Bcn-A appears to be activated upon secretion based on cell fractionation assays. The other two loci were each shown to be single ORFs encoding Bcn-B and Bcn-C. Both gene products possess homology toward known proteases. Proteinase activity for both Bcn-B and Bcn-C was confirmed using a milk agar assay. Bcn-B is predicted to be an ArgC-like serine protease, which was confirmed by PMSF inhibition of proteolytic activity, whereas Bcn-C has greater than 50% amino acid sequence identity to two zinc metalloproteases.

Influence of weather variables on spore biology of Corynespora cassiicola, an incitant of target leaf spot disease of tomato

Among the various biotic and abiotic factors limiting production of tomato, target leaf spot disease caused by Corynespora cassiicola has been emerged as a serious problem in vegetable based cropping systems. In the current research work, the influence of varying environmental conditions including temperature, light, and relative humidity on conidial germination (CG), germ tube elongation (GTE) and sporulation (SP) of C. cassiicola was studied under controlled conditions. The temperature of 25 °C was most congenial for CG and GTE both under light and darkness conditions. The temperature range of 25 °C–30 °C was optimum for CG. Conidial germination increased progressively from 15 to 25 °C both under light and dark conditions and decreased with further increase in temperature. High rate of CG was observed under continuous darkness in comparison to continuous light irrespective of temperature. Lesser duration of light had a stimulatory influence on SP of C. cassiicola and 10 h light/14 h dark cycle at 25 °C temperature was most optimum for SP. Relative humidity (RH) at 95% was optimum for CG, GTE and SP and thus, justified the peak occurrence of the target leaf spot disease during 50-4 Standard meteorological week (SMW). Comparatively higher level of CG was obtained at the humidity range of 95%–100% with the best at 100% RH (92.6%). Increasing humidity significantly decreased the time of spore production. Maximum time required for spore production (69.2 h) was observed at 85% RH followed by 95% RH (50.2 h).

Tomato bHLH132 Transcription Factor Controls Growth and Defense and Is Activated by Xanthomonas euvesicatoria Effector XopD During Pathogenesis

Effector-dependent manipulation of host transcription is a key virulence mechanism used by Xanthomonas species causing bacterial spot disease in tomato and pepper. Transcription activator-like (TAL) effectors employ novel DNA-binding domains to directly activate host transcription, whereas the non-TAL effector XopD uses a small ubiquitin-like modifier (SUMO) protease activity to represses host transcription. The targets of TAL and non-TAL effectors provide insight to the genes governing susceptibility and resistance during Xanthomonas infection. In this study, we investigated the extent to which the X. euvesicatoria non-TAL effector strain Xe85-10 activates tomato transcription to gain new insight to the transcriptional circuits and virulence mechanisms associated with Xanthomonas euvesicatoria pathogenesis. Using transcriptional profiling, we identified a putative basic helix-loop-helix (bHLH) transcription factor, bHLH132, as a pathogen-responsive gene that is moderately induced by microbe-associated molecular patterns and defense hormones and is highly induced by XopD during X. euvesicatoria infection. We also found that activation of bHLH132 transcription requires the XopD SUMO protease activity. Silencing bHLH132 mRNA expression results in stunted tomato plants with enhanced susceptibility to X. euvesicatoria infection. Our work suggests that bHLH132 is required for normal vegetative growth and development as well as resistance to X. euvesicatoria. It also suggests new transcription-based models describing XopD virulence and recognition in tomato.

Target leaf spot of tomato incited by Corynespora cassiicola, an emerging disease in tomato production under Gangetic alluvial region of West Bengal, India

Tomato (Lycopersicon esculentum Mill.) is a cosmopolitan vegetable and widely cultivated in almost all the countries of the world including India. Irreversible investment – production ratio for tomato cultivation in recent Indian agricultural systems arise the question, is there any biotic backlogs responsible for such a production loss. Our present investigation is based on this fact. Target leaf spot disease of tomato is caused by Corynespora cassiicola, a serious and emerging disease in India. Prolonged real time surveillance of this disease on tomato from 2010–11 to 2016–17, reflects some remarkable features of pathogenic progress in Gangetic alluvial region of West Bengal. This pathogen is the natural barrier for tomato production with a disease severity ranged between 35% and 58% which ultimately causes tremendous loss of tomato foliage and fruits. C. cassiicola was identified on the basis of morpho-cultural (ITCC Accession No. 7542) and molecular characterization (Genbank Accession No. KJ767193). Homology searching of internal transcribed spacer region of CcHaTom isolate was highly matched with Genbank Accession No. KP666184 (Cynodondactylon/India), AB873045 (Vitex negundo/India) and JN541214 (Malvaviscus concinnus/USA) with 95% similarity. Phylogenetic analysis established that KJ767193 and C. cassiicola retrieved sequences were conspecific from a common ancestral origin, which supports its neighbourhood with this fungal pathogen. Optimum temperature between 24 and 25 °C, coupled with 80–85% relative humidity triggered the disease progress. From the emerging scenario, C. cassiicola infecting tomato is the real threat for indigenous cultivars.

Draft Genome Sequences of Sporulating (CIDEFI-213) and Nonsporulating (CIDEFI-212) Strains of Stemphylium lycopersici.

Stemphylium lycopersici (Pleosporales) is a pathogenic fungus found on a broad range of plant hosts. It is one of the causal agents of gray leaf spot disease in tomato that causes severe yield reductions and economic losses worldwide. Here, we present the draft genome sequences and the gene annotations of two strains of S. lycopersici, nonsporulating strain CIDEFI-212 and sporulating strain CIDEFI-213.

Improvement of biomass and cyclic lipopeptides production in Bacillus amyloliquefaciens MEP218 by modifying carbon and nitrogen sources and ratios of the culture media

Bacillus amyloliquefaciens MEP218 is a native isolate with a broad spectrum of antifungal activity against plant pathogenic fungi. The ability of strain MEP218 to antagonize phytopathogens is due to the production of cyclic lipopeptides (CLPs). In this work, different carbon (C) and nitrogen (N) sources and C to N ratios were evaluated in order to improve both, biomass and CLPs production by strain MEP218. Among the C and N sources and C to N ratios tested, glucose and NH4NO3 at the C to N ratio of 10:1 enhanced significantly biomass and CLPs yield. Moreover, CLPs produced in this medium exhibited higher antibacterial activity against Xanthomonas axonopodis pv. vesicatoria (Xav) than those obtained in the recommended medium for CLPs production. Interestingly, CLPs addition influenced the development of Xav biofilm on biotic and abiotic surfaces. A comparison of HPLC-chromatograms of CLPs obtained in the optimized medium versus the ones obtained in the recommended medium showed a notable increase of surfactin in the CLPs obtained in the optimized medium. Furthermore, two peaks with antibacterial activity against Xav, identified by mass spectrometry analysis as fengycins A and B, were detected in the CLPs from the strain MEP218, grown in the optimized culture medium. The results obtained in this work suggest that changes in C and N sources and C to N ratios affect the yield and type of CLPs produced by B. amyloliquefaciens MEP218. To the best of our knowledge, this is the first study to report the finding of fengycins with antibacterial activity. CLPs produced by the strain MEP218 are potential candidates for controlling bacterial spot disease in tomato and pepper.

The mitochondrial genome of the plant-pathogenic fungus Stemphylium lycopersici uncovers a dynamic structure due to repetitive and mobile elements.

Stemphylium lycopersici (Pleosporales) is a plant-pathogenic fungus that has been associated with a broad range of plant-hosts worldwide. It is one of the causative agents of gray leaf spot disease in tomato and pepper. The aim of this work was to characterize the mitochondrial genome of S. lycopersici CIDEFI-216, to use it to trace taxonomic relationships with other fungal taxa and to get insights into the evolutionary history of this phytopathogen. The complete mitochondrial genome was assembled into a circular double-stranded DNA molecule of 75,911 bp that harbors a set of 37 protein-coding genes, 2 rRNA genes (rns and rnl) and 28 tRNA genes, which are transcribed from both sense and antisense strands. Remarkably, its gene repertoire lacks both atp8 and atp9, contains a free-standing gene for the ribosomal protein S3 (rps3) and includes 13 genes with homing endonuclease domains that are mostly located within its 15 group I introns. Strikingly, subunits 1 and 2 of cytochrome oxidase are encoded by a single continuous open reading frame (ORF). A comparative mitogenomic analysis revealed the large extent of structural rearrangements among representatives of Pleosporales, showing the plasticity of their mitochondrial genomes. Finally, an exhaustive phylogenetic analysis of the subphylum Pezizomycotina based on mitochondrial data reconstructed their relationships in concordance with several studies based on nuclear data. This is the first report of a mitochondrial genome belonging to a representative of the family Pleosporaceae.

Induction of defence-related enzymes in tomato (Solanum lycopersicum) plants treated with Bacillus subtilis CBR05 against Xanthomonas campestris pv. vesicatoria

ABSTRACTBacterial spot disease caused by Xanthomonas campestris pv. vesicatoria is one of the most important destructive diseases of tomato in many parts of the agricultural world. Therefore, the present study aims to determine the effects of Bacillus subtilis CBR05 inoculation on bacterial spot disease severity and the induction of defence-related enzymes response in tomato. Tomato leaves were evaluated to determine the activities of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and polyphenol oxidase (PPO)) and the content of malondialdehyde (MDA). A reduction in bacterial spot severity was observed in plants inoculated with B. subtilis, compared with those of uninoculated controls. A significant increase in SOD, CAT, POD, and PPO activities was observed in plants treated with B. subtilis after 24 h inoculation compared with non-inoculated pathogen control and mock-inoculated controls. Moreover, the MDA content was induced by pathogen infection, and its amount in B. subtilis inoculated plants was significantly lower than that in pathogen control. Our results suggest that early increases in antioxidant enzymes and the reduction in MDA content with B. subtilis inoculation may play a pivotal role in mitigating oxidative stress, thereby induced systemic resistance against bacterial spot disease in tomato.

Xanthomonas campestris pv. vesicatoria Secretes Proteases and Xylanases via the Xps Type II Secretion System and Outer Membrane Vesicles.

Many plant-pathogenic bacteria utilize type II secretion (T2S) systems to secrete degradative enzymes into the extracellular milieu. T2S substrates presumably mediate the degradation of plant cell wall components during the host-pathogen interaction and thus promote bacterial virulence. Previously, the Xps-T2S system from Xanthomonas campestris pv. vesicatoria was shown to contribute to extracellular protease activity and the secretion of a virulence-associated xylanase. The identities and functions of additional T2S substrates from X. campestris pv. vesicatoria, however, are still unknown. In the present study, the analysis of 25 candidate proteins from X. campestris pv. vesicatoria led to the identification of two type II secreted predicted xylanases, a putative protease and a lipase which was previously identified as a virulence factor of X. campestris pv. vesicatoria. Studies with mutant strains revealed that the identified xylanases and the protease contribute to virulence and in planta growth of X. campestris pv. vesicatoria. When analyzed in the related pathogen X. campestris pv. campestris, several T2S substrates from X. campestris pv. vesicatoria were secreted independently of the T2S systems, presumably because of differences in the T2S substrate specificities of the two pathogens. Furthermore, in X. campestris pv. vesicatoria T2S mutants, secretion of T2S substrates was not completely absent, suggesting the contribution of additional transport systems to protein secretion. In line with this hypothesis, T2S substrates were detected in outer membrane vesicles, which were frequently observed for X. campestris pv. vesicatoria. We, therefore, propose that extracellular virulence-associated enzymes from X. campestris pv. vesicatoria are targeted to the Xps-T2S system and to outer membrane vesicles. The virulence of plant-pathogenic bacteria often depends on TS2 systems, which secrete degradative enzymes into the extracellular milieu. T2S substrates are being studied in several plant-pathogenic bacteria, including Xanthomonas campestris pv. vesicatoria, which causes bacterial spot disease in tomato and pepper. Here, we show that the T2S system from X. campestris pv. vesicatoria secretes virulence-associated xylanases, a predicted protease, and a lipase. Secretion assays with the related pathogen X. campestris pv. campestris revealed important differences in the T2S substrate specificities of the two pathogens. Furthermore, electron microscopy showed that T2S substrates from X. campestris pv. vesicatoria are targeted to outer membrane vesicles (OMVs). Our results, therefore, suggest that OMVs provide an alternative transport route for type II secreted extracellular enzymes.

Functional and proteomic analyses reveal that wxcB is involved in virulence, motility, detergent tolerance, and biofilm formation in Xanthomonas campestris pv. vesicatoria

The bacterial envelope possesses diverse functions, including protection against environmental stress and virulence factors for host infection. Here, we report the function of wxcB in Xanthomonas campestris pv. vesicatoria (Xcv), a causal agent of bacterial leaf spot disease in tomato and pepper. To characterize roles of wxcB, we generated a knockout mutant (XcvΔwxcB) and found that the virulence of the mutant was weaker than that of the wild type in tomato plants. To predict the mechanism affected by wxcB, we compared protein expressions between the wild type and the mutant. Expression of 152 proteins showed a greater than 2-fold difference. Proteins involved in motility and cell wall/membrane were the most abundant. Through phenotypic assays, we further demonstrated that the mutant displayed reduced motility and tolerance to treatment, but it showed increased biofilm formation. Interestingly, the LPS profile was unchanged. These results lead to new insights into the functions of wxcB that is associated with cell wall/membrane functions, which contributes to pathogen virulence.

Transgenic resistance confers effective field level control of bacterial spot disease in tomato.

We investigated whether lines of transgenic tomato (Solanum lycopersicum) expressing the Bs2 resistance gene from pepper, a close relative of tomato, demonstrate improved resistance to bacterial spot disease caused by Xanthomonas species in replicated multi-year field trials under commercial type growing conditions. We report that the presence of the Bs2 gene in the highly susceptible VF 36 background reduced disease to extremely low levels, and VF 36-Bs2 plants displayed the lowest disease severity amongst all tomato varieties tested, including commercial and breeding lines with host resistance. Yields of marketable fruit from transgenic lines were typically 2.5 times that of the non-transformed parent line, but varied between 1.5 and 11.5 fold depending on weather conditions and disease pressure. Trials were conducted without application of any copper-based bactericides, presently in wide use despite negative impacts on the environment. This is the first demonstration of effective field resistance in a transgenic genotype based on a plant R gene and provides an opportunity for control of a devastating pathogen while eliminating ineffective copper pesticides.

Expression of Xanthomonas campestris pv. vesicatoria Type III Effectors in Yeast Affects Cell Growth and Viability

The gram-negative bacterium Xanthomonas campestris pv. vesicatoria is the causal agent of spot disease in tomato and pepper. X. campestris pv. vesicatoria pathogenicity depends on a type III secretion system delivering effector proteins into the host cells. We hypothesized that some X. campestris pv. vesicatoria effectors target conserved eukaryotic cellular processes and examined phenotypes induced by their expression in yeast. Out of 21 effectors tested, 14 inhibited yeast growth in normal or stress conditions. Viability assay revealed that XopB and XopF2 attenuated cell proliferation, while AvrRxo1, XopX, and XopE1 were cytotoxic. Inspection of morphological features and DNA content of yeast cells indicated that cytotoxicity caused by XopX and AvrRxo1 was associated with cell-cycle arrest at G0/1. Interestingly, XopB, XopE1, XopF2, XopX, and AvrRxo1 that inhibited growth in yeast also caused phenotypes, such as chlorosis and cell death, when expressed in either host or nonhost plants. Finally, the ability of several effectors to cause phenotypes in yeast and plants was dependent on their putative catalytic residues or localization motifs. This study supports the use of yeast as a heterologous system for functional analysis of X. campestris pv. vesicatoria type III effectors, and sets the stage for identification of their eukaryotic molecular targets and modes of action.

The SlMKK2 and SlMPK2 genes play a role in tomato disease resistance to Xanthomonas campestris pv. vesicatoria

Xanthomonas campestris pv. vesicatoria (Xcv) is the causal agent of bacterial spot disease in tomato (Solanum lycopersicum) plants. We recently identified a MAPKKK gene, SlMAPKKKε, which is required for tomato resistance to Xcv strains and encodes a positive regulator of cell death. We also provided evidence that the MEK2 MAPKK, and the WIPK and SIPK MAPKs act downstream to MAPKKKε in Nicotiana benthamiana plants. Here, we used the virus-induced gene silencing technique to assess whether tomato homologs of MEK2 (SlMKK2), SIPK (SlMPK1 and SlMPK2), WIPK (SlMPK3), and other components of MAP kinase cascades (SlNPK1, SlMEK1 and SlNTF6), which were previously implicated in plant immunity, are involved in disease resistance to Xcv. Silencing of none of the tested genes caused the appearance of disease symptoms in tomato leaves challenged with an avirulent Xcv strain. However, bacterial populations were significantly higher in leaves of plants silenced for SlMKK2 and SlMPK2, as compared to control plants, suggesting that these two genes contribute to disease resistance to Xcv. It remains to be established whether SlMKK2 and SlMPK2 are activated by SlMAPKKKε directly or through a distinct MAPKKK.