Bacterial Spot Disease Of Tomato Research Articles

Multiple Acquisitions of XopJ2 Effectors in Populations of Xanthomonas perforans.

Type III effectors (T3Es) are major determinants of Xanthomonas virulence and targets for resistance breeding. XopJ2 (synonym AvrBsT) is a highly conserved YopJ-family T3E acquired by X. perforans, the pathogen responsible for bacterial spot disease of tomato. In this study, we characterized a new variant (XopJ2b) of XopJ2, which is predicted to have a similar three-dimensional (3D) structure as the canonical XopJ2 (XopJ2a) despite sharing only 70% sequence identity. XopJ2b carries an acetyltransferase domain and the critical residues required for its activity, and the positions of these residues are predicted to be conserved in the 3D structure of the proteins. We demonstrated that XopJ2b is a functional T3E and triggers a hypersensitive response (HR) when translocated into pepper cells. Like XopJ2a, XopJ2b triggers HR in Arabidopsis that is suppressed by the deacetylase, SOBER1. We found xopJ2b in genome sequences of X. euvesicatoria, X. citri, X. guizotiae, and X. vasicola strains, suggesting widespread horizontal transfer. In X. perforans, xopJ2b was present in strains collected in North America, Africa, Asia, Australia, and Europe, whereas xopJ2a had a narrower geographic distribution. This study expands the Xanthomonas T3E repertoire, demonstrates functional conservation in T3E evolution, and further supports the importance of XopJ2 in X. perforans fitness on tomato. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

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.

Potential of Novel Magnesium Nanomaterials to Manage Bacterial Spot Disease of Tomato in Greenhouse and Field Conditions.

Bacterial spot of tomato is among the most economically relevant diseases affecting tomato plants globally. In previous studies, non-formulated magnesium oxide nanoparticles (nano-MgOs) significantly reduced the disease severity in greenhouse and field conditions. However, the aggregation of nano-MgO in liquid suspension makes it challenging to use in field applications. Therefore, we formulated two novel MgO nanomaterials (SgMg #3 and SgMg #2.5) and one MgOH2 nanomaterial (SgMc) and evaluated their physical characteristics, antibacterial properties, and disease reduction abilities. Among the three Mg nanomaterials, SgMc showed the highest efficacy against copper-tolerant strains of Xanthomonas perforans in vitro, and provided disease reduction in the greenhouse experiments compared with commercial Cu bactericide and an untreated control. However, SgMc was not consistently effective in field conditions. To determine the cause of its inconsistent efficacy in different environments, we monitored particle size, zeta potential, morphology, and crystallinity for all three formulated materials and nano-MgOs. The MgO particle size was determined by the scanning electron microscopy (SEM) and dynamic light scattering (DLS) techniques. An X-ray diffraction (XRD) study confirmed a change in the crystallinity of MgO from a periclase to an Mg(OH)2 brucite crystal structure. As a result, the bactericidal activity correlated with the high crystallinity present in nano-MgOs and SgMc, while the inconsistent antimicrobial potency of SgMg #3 and SgMg #2.5 might have been related to loss of crystallinity. Future studies are needed to determine which specific variables impair the performance of these nanomaterials in the field compared to under greenhouse conditions. Although SgMc did not lead to significant disease severity reduction in the field, it still has the potential to act as an alternative to Cu against bacterial spot disease in tomato transplant production.

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.

Genome-Wide Association to Study the Host-Specificity Determinants of Xanthomonas perforans.

Xanthomonas perforans and X. euvesicatoria are the causal agents of bacterial spot disease of tomato and pepper, endemic to the Southeastern United States. Although very closely related, the two bacterial species differ in host-specificity, where X. perforans is the dominant pathogen of tomato and X. euvesicatoria of pepper. This is in part due to the activity of avirulence proteins that are secreted by X. perforans strains and elicit an effector-triggered immunity (ETI) in pepper leaves, thereby restricting pathogen growth. In recent years, the emergence of several pepper-pathogenic X. perforans lineages has revealed variability within the bacterial species to multiply and cause disease in pepper, even in the absence of avirulence gene activity. Here, we investigated the basal evolutionary processes underlying the host-range of this species using multiple genome-wide association analyses. Surprisingly, we identified two novel gene-candidates that were significantly associated with pepper-pathogenic X. perforans and X. euvesicatoria. Both candidates were predicted to be involved in the transport/acquisition of nutrients common to plant-cell wall or apoplast and included a TonB-dependent receptor, which was disrupted through independent mutations within the X. perforans lineage. The other included a symporter of protons/glutamate, gltP, enriched with pepper-associated mutations near the promoter and start codon of the gene. Functional analysis of these candidates revealed that only the TonB-dependent receptor had a minor effect on the symptom development and growth of X. perforans in pepper leaves, indicating that pathogenicity to this host may have evolved independently within the bacterial species and is likely a complex, multigenic trait.

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

Antibacterial Activity of Copper Nanoparticles against Xanthomonas campestris pv. vesicatoria in Tomato Plants.

Copper-based bactericides have appeared as a new tool in crop protection and offer an effective solution to combat bacterial resistance. In this work, two copper nanoparticle products that were previously synthesized and evaluated against major bacterial and fungal pathogens were tested on their ability to control the bacterial spot disease of tomato. Growth of Xanthomonas campestris pv. vesicatoria, the causal agent of the disease, was significantly suppressed by both nanoparticles, which had superior function compared to conventional commercial formulations of copper. X-ray fluorescence spectrometry measurements in tomato leaves revealed that bioavailability of copper is superior in the case of nanoparticles compared to conventional formulations and is dependent on synthesis rather than size. This is the first report correlating bioavailability of copper to nanoparticle efficacy.

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.

Utilizing Tajima's D to identify potential microbe-associated molecular patterns in Xanthomonas euvesicatoria and X. perforans

The coevolution between plants and pathogens constitutes a significant proportion of evolutionary history for both host and parasite. Here we used the population genetics test statistic Tajima's D to identify genes involved in plant-pathogen interactions between tomato and Xanthomonas euvesicatoria and X. perforans, the causal agents of bacterial spot disease of tomato. We assessed the usefulness of this method and experimentally confirmed alleles of known microbe-associated molecular patterns (MAMPs). Overall, we demonstrated the utility of population genetic statistics such as Tajima's D for the identification of genes that may be important for pathogen recognition by the host immune machinery.

Copper-fixed quat: a hybrid nanoparticle for application as a locally systemic pesticide (LSP) to manage bacterial spot disease of tomato

The development of bacterial tolerance against pesticides poses a serious threat to the sustainability of food production. Widespread use of copper (Cu)-based products for plant disease management has led to the emergence of copper-tolerant pathogens such as Xanthomonas perforans (X. perforans) strains in Florida, which is very destructive to the tomato (Solanum lycopersicum) industry. In this study, we report a hybrid nanoparticle (NP)-based system, coined Locally Systemic Pesticide (LSP), which has been designed for improved efficacy compared to conventional Cu-based bactericides against Cu-tolerant X. perforans. The silica core–shell structure of LSP particles makes it possible to host ultra-small Cu NPs (<10 nm) and quaternary ammonium (Quat) molecules on the shell. The morphology, release of Cu and Quat, and subsequent in vitro antimicrobial properties were characterized for LSP NPs with core diameters from 50 to 600 nm. A concentration of 4 μg mL−1 (Cu): 1 μg mL−1 (Quat) was found to be sufficient to inhibit the growth of Cu-tolerant X. perforans compared to 100 μg mL−1 (metallic Cu) required with standard Kocide 3000. Wetting properties of LSP exhibited contact angles below 60°, which constitutes a significant improvement from the 90° and 85° observed with water and Kocide 3000, respectively. The design was also found to provide slow Cu release to the leaves upon water washes, and to mitigate the phytotoxicity of water-soluble Cu and Quat agents. With Cu and Quat bound to the LSP silica core–shell structure, no sign of phytotoxicity was observed even at 1000 μg mL−1 (Cu). In greenhouse and field experiments, LSP formulations significantly reduced the severity of bacterial spot disease compared to the water control. Overall, the study highlights the potential of using LSP particles as a candidate for managing tomato bacterial spot disease and beyond.

Plant Extract Treatments Induce Resistance to Bacterial Spot by Tomato Plants for a Sustainable System

The aim of this study is to assess the effect of extracts of Nerium oleander, Eucalyptus chamadulonsis and Citrullus colocynthis against bacterial spot disease of tomato and to investigate the induction of resistance by tomato (Solanum lycopersicum) in order to promote a sustainable management system. The antibacterial activity of aqueous and ethanol plant extracts was tested against Xanthomonas axonopodis pv. vesicatoria, isolate PHYXV3, in vitro and in vivo. The highest antibacterial activity in vitro was obtained with C. colocynthis, N. oleander and E. chamadulonsis, respectively. In vivo, ethanol extracts of N. oleander and E. chamadulonsis were more effective than aqueous extracts in reducing pathogen populations on tomato leaves. Under greenhouse conditions, application of the plant extracts at 15% (v/v) to tomato plants significantly reduced disease severity and increased the shoot weight of ‘Super Marmande’ tomato. In most cases, plant extracts significantly increased total phenol and salicylic acid content of tomato plants compared to either healthy or infected ones. In addition, C. colocynthis and E. chamadulonsis extracts significantly increased peroxidase activity while only E. chamadulonsis increased polyphenol oxidase after infection with the causal agent. The results indicated that the plant extracts showed promising antibacterial activity and could be considered an effective tool in integrated management programs for a sustainable system of tomato bacterial spot control.

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.

Nanoscale quantification of longitudinal and transverse mechanics of bacterial bodies

Mechanical studies focused on flagella and pili of bacteria, typically estimated by evaluating bending persistence length measurements or Young's modulus, fail to consider the behavior of the whole bacterial body, which can undergo significant changes during antimicrobial resistance. In this Letter, we investigate the biophysical properties of an important plant bacterial pathogen, Xanthomonas perforans, the causal agent for bacterial spot disease of tomato and pepper. The emergence of copper-tolerant strains of X. perforans is predicted to have a devastating impact on industrial scale productions, making it essential to advance approaches to distinguish copper-tolerant from the copper-sensitive X. perforans strains. After introducing a means to measure the bending stiffness of bacteria using atomic force microscopy (AFM), we propose a comparison of the longitudinal and transverse mechanical properties by acquiring high resolution images and force curves of the bacteria. A quantification of the longitudinal and transverse Young's modulus of bacteria reveals significant differences between copper-sensitive and copper-tolerant strains of X. perforans. The present approach is expected to significantly advance the fundamental understanding of bacterial phenomena, such as by monitoring the therapeutic efficacy of newly developed materials on individual X. perforans in situ to relate them to the behavior of their colonies or biofilms.

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.

Effectiveness of tailocins produced by Pseudomonas fluorescens SF4c in controlling the bacterial-spot disease in tomatoes caused by Xanthomonas vesicatoria

The development of alternatives for the use of chemical pesticides for plant disease control is the present-day and ongoing challenge for achieving sustainable agriculture. Pseudomonas fluorescens SF4c, native strain from wheat, produces tailocins (phage-tail-like bacteriocins) with antimicrobial activity against several phytopathogenic strains. We thus investigated the efficacy of foliar application of these bacteriocins to control the bacterial-spot disease in tomato caused by Xanthomonas vesicatoria Xcv Bv5-4a. The disease severity and incidence index were reduced by 44 and 36%, respectively; while the number of viable cells of X. vesicatoria Xcv Bv5-4a decreased after bacteriocin treatment. Furthermore, bacteriocin was effective in reducing bacterial-spot-disease symptoms on tomato fruits even when applied 12 h after infection. Tailocin activity was not affected by abiotic influences such as adjuvant, light and temperature and, biotic factors such as apoplastic-fluids. In contrast, no antibacterial activity of these tailocins was observed when the bacteriocin was exposed to extremely dry conditions. Finally, that no cytotoxic effects on mammalian cells were observed with this representative tailocins is highly significant and demonstrates the safety of such compounds in humans. All these findings indicate that the SF4c tailocins represent an attractive alternative to copper-containing bactericides for use in the control of bacterial spot.

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.

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.

2,6-Dichloroisonicotinic acid enhances the expression of defense genes in tomato seedlings against Xanthomonas perforans

Plant activation is an appealing disease management tool, avoiding some of the challenges of traditional chemical control by not directly impacting the pathogen. 2,6-dichloroisonicotinic acid (INA) was used as abiotic resistance inducer against bacterial spot disease of tomato caused by Xanthomonas perforans. Activities of Ascorbate peroxidase (APX) and Catalase (CAT), increased in INA primed tomato seedlings, which were challenged with X. perforans and enzyme activities of APX and CAT reached maximum at 9 hpi and 15 hpi, in resistant cultivar whereas in highly susceptible they took 18 hpi and 24 hpi respectively. The transcripts accumulation studies for APX2 and CAT1 were carried out using semiquantitative PCR, with maximum mRNA accumulation in resistant cultivar upon INA priming. Quantitative real time-PCR (qRT-PCR) confirmed the maximum induction of both the genes in INA primed seedlings. Seed priming with 1.0 mM INA effectively reduced disease severity in tomato plants under field conditions. Moreover, these enzyme activities were significantly increased in INA primed and X. perforans challenged tomato seedlings. Expression of APX2 and CAT1 genes, in tomato were rapidly and significantly up-regulated by INA priming. Furthermore, a subsequent challenge of INA primed seedlings with X. perforans resulted in higher expression of defense genes. However, the expression of defense genes was more in INA primed resistant cultivar than highly susceptible cultivar. Therefore, our results suggest that seed priming with INA could enhance disease protection against X. perforans infection in tomato through the activation of defense genes.

Photocatalysis: Effect of Light-Activated Nanoscale Formulations of TiO2 on Xanthomonas perforans and Control of Bacterial Spot of Tomato

Protection of crops from bacterial diseases presents a continuing challenge, mandating the development of novel agents and approaches. Photocatalysis is a process where chemically reactive oxygen species are catalytically generated by certain minerals in the presence of light. These reactive oxygen species have the capacity to destroy organic molecular structures critical to pathogen viability. In this study, the antibacterial potential of photocatalytic nanoscale titanium dioxide (TiO(2)), nanoscale TiO(2) doped (incorporation of other materials into the structure of TiO(2)) with silver (TiO(2)/Ag), and nanoscale TiO(2) doped with zinc (TiO(2)/Zn; AgriTitan) was evaluated against Xanthomonas perforans, the causal agent for bacterial spot disease of tomato. In vitro experiments on photocatalytic activity and dose dependency were conducted on glass cover slips coated with the nanoscale formulations by adding a known population of X. perforans strain Xp-F7 and illuminating the cover slips under a visible light source. TiO(2)/Ag and TiO(2)/Zn had high photocatalytic activity against X. perforans within 10 min of exposure to 3 × 10(4) lux. Greenhouse studies on naturally and artificially infected transplants treated with TiO(2)/Zn at ≈500 to 800 ppm significantly reduced bacterial spot severity compared with untreated and copper control. Protection was similar to the grower standard, copper + mancozeb. The use of TiO(2)/Zn at ≈500 to 800 ppm significantly reduced disease incidence in three of the four trials compared with untreated and copper control, and was comparable to or better than the grower standard. The treatments did not cause any adverse effects on tomato yield in any of the field trials.