Susceptible Tomato Varieties Research Articles

Genetic diversity and heritability of tomato parental lines assembled for Ralstonia solanacearum resistance

Bacterial wilt is a disease caused by Ralstonia solanacearum, which affects over 450 plant species and causes significant reduction in crop yields including of tomato (Solanum lycopersicum) worldwide. Developing and identifying tomato genotypes with Ralstonia solanacearum tolerance and high yield potential, presents an opportunity for improvement of crop productivity. The objective of this study was to explore the heritability of tomato resistance against bacterial wilt and the genetic variation within the population for breeding purposes. A breeding population was created by crossing two bacterial wilt-resistant (MT56 and BL333) and three commercially desirable susceptible (Assila, Rambo and Heinz) tomato varieties, using North Carolina Design II, in a screen house at the Makerere University Agricultural Research Institute, Kabanyolo (MUARIK). Results showed that the Area Under Disease Progress Curve was significant (P<0.001), indicating that cumulative disease progress was less in resistant genotypes of segregating generations. Disease severity increased with days after inoculation (DAI), with Heinz showing the highest level of susceptibility. The General Combining Ability for male parent (GCAm) was significant (P< 0.01), and Specific Combining Ability (SCAf×m) and GCAf were significant (P<0.01) for the F2 generation. Broad-sense heritability was higher than the narrow-sense heritability in both F1 and F2 generations, suggesting that non-additive gene action predominately controlled tomato resistance to bacterial wilt infestation. The genetic diversity ranged from 0.5 to 0.6759, with a mean value of 0.5787. Polymorphism Information Content (PIC) varied from 0.375 to 0.6357, with a mean of 0.4888, indicating a high degree of variation. SLM 12-2 was the most polymorphic marker, with a PIC of 0.6357. The Unweighted Pair-Group Method with Arithmetic Average (UPGMA) classified all tomato genotypes into six clusters, namely Clusters 1 and 2 (Susceptible parents), Cluster 3 (Resistant parents), Clusters 4 and 5 (New source of resistance), and Cluster 6 (F1P1×P5).

Dicer-Like Protein 4 and RNA-Dependent RNA Polymerase 6 Are Involved in Tomato Torrado Virus Pathogenesis in Nicotiana benthamiana.

Tomato torrado virus (ToTV) is a type member of the Torradovirus genus in the Secoviridae family known to cause severe necrosis in susceptible tomato varieties. ToTV also infects other Solanaceae plants, including Nicotiana benthamiana, where it induces distinctive disease symptoms: plant growth drop with the emergence of spoon-like malformed systemic leaves. Virus-induced post-transcriptional gene silencing (PTGS) is significant among plant defense mechanisms activated upon virus invasion. The PTGS, however, can be counteracted by suppressors of RNA silencing commonly found in viruses, which efficiently disrupt the antiviral defense of their host. Here, we addressed the question of PTGS antiviral activity and its suppression in N. benthamiana during ToTV infection-a phenomenon not described for any representative from the Torradovirus genus so far. First, we showed that neither the Vp26-a necrosis-inducing pathogenicity determinant of ToTV-nor other structural viral proteins limited the locally induced PTGS similar to p19, a well-characterized potent suppressor of RNA silencing of tombusviruses. Moreover, by employing wild-type and transgenic lines of N. benthamiana with suppressed Dicer-like 2 (DCL2), Dicer-like 4 (DCL4), Argonaute 2 and RNA-dependent RNA polymerase 6 (RDR6) proteins, we proved their involvement in anti-ToTV defense. Additionally, we identified DCL4 as the major processor of ToTV-derived siRNA. More importantly, our results indicate the essential role of the Suppressor of Gene Silencing 3 (SGS3)/RDR6 pathway in anti-ToTV defense. Finally, we conclude that ToTV might not require a potent RNA silencing suppressor during infection of the model plant N. benthamiana.

Contribuição do QuitoMax® no metabolismo hormonal e enzimático em tomate sob estresse salino

ABSTRACT Salinity stress severely restricts plant nutrition and hinders biochemical and physiological processes crucial for growth. In several crop systems bioactive products which confer growth promotion, are applied as a sustainable alternative for contributing to food security. The aim of this work was to evaluate the biochemical contribution of QuitoMax® to hormonal and enzymatic metabolism in tomato under saline stress. Three treatments were applied: saline without QuitoMax®, nonsaline + QuitoMax® and saline + QuitoMax®. A tolerant (Amalia) and a susceptible (Claudia) tomato variety were used as experimental models. The normalized difference vegetation index (NDVI) was measured as a morphological variable, and peroxidase (POD), glutamine synthetase (GS) and nitrate reductase (NR) enzyme activities were determined. Gibberellic (GA) and abscisic acid (ABA) concentrations were also determined. Due to the effects of QuitoMax®, the plants maintained high NDVI values even under saline conditions. A decrease in POD and GS activity and an increase in NR activity were also found. The GA concentration in the leaves was higher in the tolerant variety when QuitoMax® was applied than in the saline treatment but lower in the susceptible variety. The opposite behavior was found when the ABA concentration was quantified. This study demonstrates the protective action of QuitoMax® under salinity stress on tomato crops in both tolerant and susceptible varieties. In crux, QuitoMax® can be opted as a shotgun approach to tackle salinity in tomato.

Metabolomics of early blight (Alternaria solani) susceptible tomato (Solanum lycopersicum) unfolds key biomarker metabolites and involved metabolic pathways

Tomato (Solanum lycopersicum) is among the most important commercial horticultural crops worldwide. The crop quality and production is largely hampered due to the fungal pathogen Alternaria solani causing necrotrophic foliage early blight disease. Crop plants usually respond to the biotic challenges with altered metabolic composition and physiological perturbations. We have deciphered altered metabolite composition, modulated metabolic pathways and identified metabolite biomarkers in A. solani-challenged susceptible tomato variety Kashi Aman using Liquid Chromatography-Mass Spectrometry (LC–MS) based metabolomics. Alteration in the metabolite feature composition of pathogen-challenged (m/z 9405) and non-challenged (m/z 9667) plant leaves including 8487 infection-exclusive and 8742 non-infection exclusive features was observed. Functional annotation revealed putatively annotated metabolites and pathway mapping indicated their enrichment in metabolic pathways, biosynthesis of secondary metabolites, ubiquinone and terpenoid-quinones, brassinosteroids, steroids, terpenoids, phenylpropanoids, carotenoids, oxy/sphingolipids and metabolism of biotin and porphyrin. PCA, multivariate PLS-DA and OPLS-DA analysis showed sample discrimination. Significantly up regulated 481 and down regulated 548 metabolite features were identified based on the fold change (threshold ≥ 2.0). OPLS-DA model based on variable importance in projection (VIP scores) and FC threshold (> 2.0) revealed 41 up regulated discriminant metabolite features annotated as sphingosine, fecosterol, melatonin, serotonin, glucose 6-phosphate, zeatin, dihydrozeatin and zeatin-β-d-glucoside. Similarly, 23 down regulated discriminant metabolites included histidinol, 4-aminobutyraldehyde, propanoate, tyramine and linalool. Melatonin and serotonin in the leaves were the two indoleamines being reported for the first time in tomato in response to the early blight pathogen. Receiver operating characteristic (ROC)-based biomarker analysis identified apigenin-7-glucoside, uridine, adenosyl-homocysteine, cGMP, tyrosine, pantothenic acid, riboflavin (as up regulated) and adenosine, homocyctine and azmaline (as down regulated) biomarkers. These results could aid in the development of metabolite-quantitative trait loci (mQTL). Furthermore, stress-induced biosynthetic pathways may be the potential targets for modifications through breeding programs or genetic engineering for improving crop performance in the fields.

Farmer knowledge, management practices and occurrence of tomato root knot nematodes in Kenya

Root-knot nematodes (RKNs) (Meloidogyne spp.) is a major constraint to tomato (Solanum lycopersicum L.) production in sub-Saharan Africa; yet there is low adoption of recommended nematode control measures in regions like east Africa due to inadequate farmer knowledge about the pest and associated diseases. The objective of this study was to assess farmers’ knowledge status, and intensity of damage of root-knot nematodes and their management practices in Kenya. A survey was conducted in Kenya, during February to August 2021; using a semi-structured questionnaire administered to 282 randomly selected household heads of actively growing tomato farmers, at two elevations in three counties. Most farmers (98.9%) could not identify the disease precisely. They mostly attributed its symptoms to moisture stress and nutrient deûciencies. Most farmers (63%) practiced mono-cropping; while only 4.3% of farmers amended soils with manure. Respondents preferred growing RKN susceptible tomato varieties, mainly Rio-Grande, Cal J, Onyx and Kilele FI. A total of 92% expressed willingness to shift to RKNs resistant varieties, if recommended varieties match their desirable characteristics. Majority of the respondents lacked knowledge on nematode characteristics and its associated disease control. A total of 37.9% of the respondents did not use control measures against the RKNs. There was high disease incidence, severity and galling index in the surveyed areas.

Grafting tomato with resistant eggplant and bacteriophages treatment to suppress the development of bacterial wilt disease (Ralstonia pseudosolanacearum)

Bacterial wilt disease caused by Ralstonia pseudosolanacearum is a major problem in tomato cultivation. This research was aimed to evaluate the potential of grafting two susceptible tomato varieties (Servo and Kaliurang) with EG203-resistant eggplant, combined with bacteriophage treatment to suppress bacterial wilt disease. The results showed that grafting tomatoes with EG203 eggplant combined with bacteriophage treatment effectively suppressed the development of bacterial wilt disease. Grafting Servo – EG203 combined with bacteriophages treatment resulted in the longest incubation period (45 days), lowest incidence (6.25%), lowest severity of bacterial wilt (2.5%), and AUDPC value (8.75) at 49 days after inoculation. The low development of bacterial wilt in grafting and bacteriophage treatment was supported by the low population of R. pseudosolanacearum in the soil, which was only around 103 CFU/gram at 49 days after inoculation. In conclusion, the combination of tomato grafting and bacteriophage treatment showed promising results in controlling bacterial wilt disease.

Comparative Transcriptome and Widely Targeted Metabolome Analysis Reveals the Molecular Mechanism of Powdery Mildew Resistance in Tomato.

Powdery mildew is a serious problem in tomato production; therefore, the PM-resistant tomato inbred line, '63187', and the susceptible tomato variety, 'Moneymaker (MM)', were used as experimental materials for the combined analysis of transcriptome and widely targeted metabolome on tomato leaves at 0 h post inoculation (hpi), 12 hpi, and 48 hpi. The results indicated that 276 genes were expressed in all treatments, and the K-means cluster analysis showed that these genes were divided into eight classes in '63187' and ten classes in 'MM'. KEGG enrichment showed that amino acid metabolism, signal transduction, energy metabolism, and other secondary metabolites biosynthesis pathways were significantly enriched. Interestingly, the analysis of WRKY family transcription factors (TFs) showed that the expression of four TFs in '63187' increased with no obvious change in 'MM'; and the expression of one TF in 'MM' increased with no obvious change in '63187'. The combined analysis revealed that both phenylpropanoid biosynthesis and flavonoid biosynthesis pathways were enriched in '63187' and 'MM'. In '63187', six metabolites involved in this pathway were downregulated, and four genes were highly expressed, while in 'MM', three metabolites were upregulated, four metabolites were downregulated, and ten genes were highly expressed. These metabolites and genes might be candidates for PM resistance or susceptibility in subsequent studies. These results provide favorable molecular information for the study of the different resistances of tomatoes to PM, and they provide a basis for the breeding of tomato varieties resistant to PM.

Pathogenicity of Alternaria Species Causing Tomato Blight in Agro-Ecological Zones of Kirinyaga County in Kenya

Tomato (Lycopersicon esculentum Mill) is a highly consumed and produced vegetable crop with an annual global production of over 185 million tonnes. High consumption and production are due to its nutritional and health benefits. However, tomato production is characterized by inconsistent quality and yields, partly emanating from biotic constraints attributed to fungal foliar diseases. In Kirinyaga County, where tomato production is a significant source of livelihood, knowledge gaps exist regarding the identity of Alternaria species causing tomato blight in different Agro-ecological zones (AEZs) and the susceptibility of locally grown tomato varieties to the Alternaria pathogens. To address these gaps, we conducted a study to determine the characteristics of an Alternaria pathogen isolated from tomato leaves from different AEZs and assess the susceptibility level of some tomato varieties grown in Kirinyaga County. Pathogen characterization was done using phenotypic and molecular approaches, and the susceptibility assay was evaluated by artificial inoculation. The molecular characterization involved BLASTIN analysis of Sanger sequenced DNA nucleotide amplicons, and the use of NCBI databases. The susceptibility assay was carried out in the greenhouse using a completely randomized design. Data on the fungal conidial lengths (µm) and the susceptibility (%) of tomato varieties to fungal isolates were subjected to Analysis of Variance (ANOVA) with significance means separated using LSD at α = 0.05. The morphologies of the pathogens differed significantly (p <0.05). Molecular characterization implicated Alternaria solani, Alternaria cerealis, Alternaria arborescens, and Alternaria alternate in causing tomato blight. Tomato varieties differed in their susceptibility to Alternaria pathogens (p < 0.05). Kilele F1 (73.84%), and Terminator F1 (56.48%) were the most and least susceptible tomato varieties, respectively. Thus, Terminator F1 can serve as a source for the genetic improvement of tomato varieties in the Kirinyaga patho-system. Our study provides valuable information on the identification of Alternaria species causing tomato blight in different AEZs in Kirinyaga County and the susceptibility of locally grown tomato varieties to infection by Alternaria pathogen isolates. The findings can aid in the development of effective disease management strategies and the genetic improvement of tomato varieties in the area.

Characterization of volatile organic compounds in grafted tomato plants upon potyvirus necrotic infection

A headspace solid-phase microextraction-gas chromatography-mass spectrometry (HS-SPME/GC-MS) method was used to study the volatile organic compounds (VOCs) associated with the differential immune response of tomato plants infected with the recombinant strain of potato virus Y (PVYC-to), necrogenic to tomato. Analysis was carried out in UC82 (UC), a virus susceptible tomato variety, comparing the same UC plants grafted or not onto a virus tolerant tomato ecotype, Manduria (Ma); the three types of samples used for the GC-MS analysis were mock-inoculated UC/Ma plants, UC/Ma+PVYC-to and UC+PVYC-to plants; the VOCs obtained were 111. Results from symptomatic PVYC-to-infected UC plants showed a VOCs composition enriched in alcohols, fatty acid derivates, benzenoids, and salicylic acid derivatives, while in mock-inoculated UC/Ma plants VOCs were mainly characterized by methyl ester compounds. The VOC profile was in line with RNAseq data analyses, denoting that PVYC-to viral RNA accumulation and disease symptoms induce the specific transcriptional activation of genes involved in VOCs biosynthesis. Furthermore, principal component analysis highlighted that VOCs of PVYC-to-infected and mock-inoculated grafted plants were much closer each other than that of symptomatic PVYC-to-infected non-grafted UC plants. These results suggest that VOCs profiles of tomato plants are related to the viral RNA accumulation, disease intensity and graft-derived tolerance to PVYC-to infection.

Effect of foliar-applied Euphorbia hirta towards controlling bacterial diseases in tomato crops and enhancing fruit yield and shelf life

ABSTRACT Bacterial diseases are a major cause of yield loss in tomato plants worldwide, and there is a need to identify strategies for control of these diseases. Bacterial wilt caused by Ralstonia solanacearum (RS) and bacterial spot caused by Xanthomonas axonopodis pv vesicatoria (Xav) are amongst the bacterial diseases affecting tomato crops. The current study investigates Euphorbia hirta (E. hirta) methanol extract (EHM) for its antibacterial activity against RS, and Xav induced diseases in tomato plants, along with its biostimulant potential in increasing fruit yield. EHM at 1280 mg/L exhibited 90% inhibition in the growth of both RS and Xav bacteria. Further, evaluation of EHM on Arka Vikas, a susceptible tomato variety, showed that pre-treatment of 15 days and 45 days tomato plants with 1280 mg/L EHM exhibited moderate resistance to bacterial wilt and spot disease, respectively, with increased tomato yield and improved chlorophyll b levels compared to control plants. The effectiveness of EHM in increasing fruit yield was further confirmed using field evaluation on ‘Namdhari NS 538’ variety of tomato plants. A 3.5-fold increase in numbers and weight of tomato was observed with foliar application of EHM (1280 mg/L). A significant reduction in fruit spoilage along with an increase in mRNA level of ethylene receptor factor (ERF1) was also observed. The study is the first to demonstrate the cost-effective and eco-friendly antibacterial and biostimulant potential of EHM (1280 mg/L) for tomato plants.

Aplikasi Actinomycetes dan Bakteriofag pada Tomat Sambung untuk Mengendalikan Penyakit Layu Bakteri Ralstonia solanacearum dan Meningkatkan Hasil Buah

Ralstonia solanacearum (Smith) is a soil-borne pathogen that causes bacterial wilt disease and is a complex species in races, biovars, phylotypes, and strains of various pathogenicities. As a result, the pathogen is difficult to control. An alternative control is by applying actinomycetes and bacteriophages on tomato grafted with resistant rootstock. This study aims to find the best combination in controlling bacterial wilt disease among grafted tomato plants and/or actinomycetes and/or bacteriophage treatments to increase yields. The graftings were between Amelia (East West, Indonesia) or H7996 (AVRDC) as rootstocks, and Servo (East West, Indonesia) as the susceptible scion. The grafting used the tube method, and the experimental design was a randomized completely block design with the grafted plants treated by actinomycetes and/or bacteriophages with three replications. The results indicated that actinomycetes application on plants grafted with Amelia rootstock or H7996 and the application of bacteriophages on susceptible varieties (Servo) could be the best alternative treatment in controlling the bacterial wilt disease. The susceptible varieties with the actinomycetes or bacteriophages application could even increase fruit yields higher than the grafted tomatoes treated with actinomycetes or bacteriophages. In fact, the susceptible tomato varieties with bacteriophage treatment showed high fruit yield.
 
 Keywords: actinomycetes, bacteriophages, bacterial wilt disease, fruit yield, Ralstonia solanacearum

Computational identification and characterization of vascular wilt pathogen (Fusarium oxysporum f. sp. lycopersici) CAZymes in tomato xylem sap

Fusarium oxysporum f. sp. lycopersici is a devastating plant pathogenic fungi known for wilt disease in the tomato plant and secrete cell wall degrading enzymes. These enzymes are collectively known as carbohydrate-active enzymes (CAZymes), crucial for growth, colonization and pathogenesis. Therefore, the present study was aimed to identify and annotate pathogen CAZymes in the xylem sap of a susceptible tomato variety using downstream proteomics and meta servers. Further, structural elucidation and conformational stability analysis of the selected CAZyme families were done through homology modeling and molecular dynamics simulation. Among all the fungal proteins identified, the carbohydrate metabolic process was found to be enriched. Most of the annotated CAZymes belonged to the hydrolase and oxidoreductase families, and 90% were soluble and extracellular. Moreover, using a publically available interactome database, interactions were observed between the families acting on chitin, hemicellulose and pectin. Subsequently, important catalytic residues were identified in the candidate CAZymes belonging to carbohydrate esterase (CE8) and glycosyl hydrolase (GH18 and GH28). Further, essential dynamics after molecular simulation of 100 ns revealed the overall behavior of these CAZymes with distinct global minima and transition states in CE8. Thus, our study identified some of the CAZyme families that assist in pathogenesis and growth through host cell wall deconstruction with further structural insight into the selected CAZyme families. Communicated by Ramaswamy H. Sarma

The Role of Grafting in the Resistance of Tomato to Viruses.

Grafting is routinely implemented in modern agriculture to manage soilborne pathogens such as fungi, oomycetes, bacteria, and viruses of solanaceous crops in a sustainable and environmentally friendly approach. Some rootstock/scion combinations use specific genetic resistance mechanisms to impact also some foliar and airborne pathogens, including arthropod or contact-transmitted viruses. These approaches resulted in poor efficiency in the management of plant viruses with superior virulence such as the strains of tomato spotted wilt virus breaking the Sw5 resistance, strains of cucumber mosaic virus carrying necrogenic satellite RNAs, and necrogenic strains of potato virus Y. Three different studies from our lab documented that suitable levels of resistance/tolerance can be obtained by grafting commercial tomato varieties onto the tomato ecotype Manduria (Ma) rescued in the framework of an Apulian (southern Italy) regional program on biodiversity. Here we review the main approaches, methods, and results of the three case studies and propose some mechanisms leading to the tolerance/resistance observed in susceptible tomato varieties grafted onto Ma as well as in self-grafted plants. The proposed mechanisms include virus movement in plants, RNA interference, genes involved in graft wound response, resilience, and tolerance to virus infection.

Grafting alters tomato transcriptome and enhances tolerance to an airborne virus infection

Grafting of commercial tomato varieties and hybrids on the tomato ecotype Manduria resulted in high levels of tolerance to the infection of Sw5 resistance-breaking strains of tomato spotted wilt virus and of severe cucumber mosaic virus strains supporting hypervirulent satellite RNAs that co-determine stunting and necrotic phenotypes in tomato. To decipher the basis of such tolerance, here we used a RNAseq analysis to study the transcriptome profiles of the Manduria ecotype and of the susceptible variety UC82, and of their graft combinations, exposed or not to infection of the potato virus Y recombinant strain PVYC-to. The analysis identified graft- and virus-responsive mRNAs differentially expressed in UC82 and Manduria, which led to an overall suitable level of tolerance to viral infection confirmed by the appearance of a recovery phenotype in Manduria and in all graft combinations. The transcriptome analysis suggested that graft wounding and viral infection had diverging effects on tomato transcriptome and that the Manduria ecotype was less responsive than the UC82 to both graft wounding and potyviral infection. We propose that the differential response to the two types of stress could account for the tolerance to viral infection observed in the Manduria ecotype as well as in the susceptible tomato variety UC82 self-grafted or grafted on the Manduria ecotype.

Development of a New Molecular Marker for the Resistance to Tomato Yellow Leaf Curl Virus.

Tomato yellow leaf curl virus (TYLCV) responsible for tomato yellow leaf curl disease (TYLCD) causes a substantial decrease in tomato (Solanum lycopersicum L.) yield worldwide. The use of resistant variety as a sustainable management strategy has been advocated. Tremendous progress has been made in genetically characterizing the resistance genes (R gene) in tomato. Breeding tomato for TYLCV resistance has been based mostly on Ty-3 as a race-specific resistance gene by introgression originating from wild tomato species relatives. Improvement or development of a cultivar is achievable through the use of marker-assisted selection (MAS). Therefore, precise and easy use of gene-targeted markers would be of significant importance for selection in breeding programs. The present study was undertaken to develop a new marker based on Ty-3 gene sequence that can be used for MAS in TYLCV resistant tomato breeding program. The new developed marker was named ACY. The reliability and accuracy of ACY were evaluated against those of Ty-3 linked marker P6-25 through screening of commercial resistant and susceptible tomato hybrids, and genetic segregation using F2 population derived from a commercial resistant hybrid AG208. With the use of bioinformatics and DNA sequencing analysis tools, deletion of 10 nucleotides was observed in Ty-3 gene sequence for susceptible tomato variety. ACY is a co-dominant indel-based marker that produced clear and strong polymorphic band patterns for resistant plant distinguishing it from its susceptible counterpart. The obtained result correlates with 3:1 segregation ratio of single resistant dominant gene inheritance, which depicted ACY as gene-tag functional marker. This marker is currently in use for screening 968 hybrids varieties and one thousand breeding lines of tomato varieties stocked in Jiangsu Green Port Modern Agriculture Development Company (Green Port). So far, ACY has been used to identify 56 hybrids and 51 breeding lines. These newly detected breeding lines were regarded as potential source of resistance for tomato breeding. This work exploited the sequence of Ty-3 and subsequently contributed to the development of molecular marker ACY to aid phenotypic selection. We thus recommend this marker to breeders, which is suitable for marker-assisted selection in tomato.

Protease Activities Triggered by Ralstonia solanacearum Infection in Susceptible and Tolerant Tomato Lines

Activity-based protein profiling (ABPP) is a powerful proteomic technique to display protein activities in a proteome. It is based on the use of small molecular probes that react with the active site of proteins in an activity-dependent manner. We used ABPP to dissect the protein activity changes that occur in the intercellular spaces of tolerant (Hawaii 7996) and susceptible (Marmande) tomato plants in response to R. solanacearum, the causing agent of bacterial wilt, one of the most destructive bacterial diseases in plants. The intercellular space -or apoplast- is the first battlefield where the plant faces R. solanacearum Here, we explore the possibility that the limited R. solanacearum colonization reported in the apoplast of tolerant tomato is partly determined by its active proteome. Our work reveals specific activation of papain-like cysteine proteases (PLCPs) and serine hydrolases (SHs) in the leaf apoplast of the tolerant tomato Hawaii 7996 on R. solanacearum infection. The P69 family members P69C and P69F, and an unannotated lipase (Solyc02g077110.2.1), were found to be post-translationally activated. In addition, protein network analysis showed that deeper changes in network topology take place in the susceptible tomato variety, suggesting that the tolerant cultivar might be more prepared to face R. solanacearum in its basal state. Altogether this work identifies significant changes in the activity of 4 PLCPs and 27 SHs in the tomato leaf apoplast in response to R. solanacearum, most of which are yet to be characterized. Our findings denote the importance of novel proteomic approaches such as ABPP to provide new insights on old and elusive questions regarding the molecular basis of resistance to R. solanacearum.

Isolation, characterization and toxicological potential of Alternaria-mycotoxins (TeA, AOH and AME) in different Alternaria species from various regions of India

Alternaria species produce various sorts of toxic metabolites during their active growth and causes severe diseases in many plants by limiting their productivity. These toxic metabolites incorporate various mycotoxins comprising of dibenzo-α-pyrone and some tetramic acid derivatives. In this study, we have screened out total 48 isolates of Alternaria from different plants belonging to different locations in India, on the basis of their pathogenic nature. Pathogenicity testing of these 48 strains on susceptible tomato variety (CO-3) showed 27.08% of the strains were highly pathogenic, 35.41% moderately pathogenic and 37.5% were less pathogenic. Phylogenetic analysis showed the presence of at least eight evolutionary cluster of the pathogen. Toxins (TeA, AOH and AME) were isolated, purified on the basis of column chromatography and TLC, and further confirmed by the HPLC-UV chromatograms using standards. The final detection of toxins was done by the LC-MS/MS analysis by their mass/charge ratio. The present study develops an approach to classify the toxicogenic effect of each of the individual mycotoxins on tomato plant and focuses their differential susceptibility to develop disease symptoms. This study represents the report of the natural occurrence and distribution of Alternaria toxins in various plants from India.

In silico prediction of miRNAs targeting ToLCV and their regulation in susceptible and resistant tomato plants

Tomato leaf curl disease is one of the main constraints in tomato production worldwide. It is induced by viruses including Tomato leaf curl virus (ToLCV). Plant viruses can both induce and be themselves targeted by gene silencing machinery. Certain animal viruses were found to be targeted by host miRNAs but, there is no similar experimental report for plant viruses. In this study, we investigated if tomato-encoded miRNAs target ToLCV using an in silico analysis. Results showed that ToLCV can be targeted by three tomato miRNAs, named miR156, miR159, and miR403. Following virus infection of susceptible tomato plants cv Moneymaker, miR159 and miR403 were upregulated, while miR156 was unchanged. In the plants of the resistant tomato variety PS550, that accumulated 50-fold less viral DNA and did not show disease symptoms, miR156 was upregulated, while the two others remained unchanged. In addition, the expression level of three genes known as targets of these miRNAs, i.e. SPL6b, MYB33 and AGO2a, was measured in both susceptible and resistant tomato varieties by qRT-PCR. The possible role of the predicted miRNAs and regulation of their target genes in symptom development by ToLCV is discussed.

Foliar Applications of Acibenzolar-S-Methyl Negatively Affect the Yield of Grafted Tomatoes in Fields Infested with Ralstonia solanacearum.

Three field experiments were conducted in Florida from 2012-2014 to assess the impact of acibenzolar-S-methyl (ASM), a systemic acquired-resistance inducer, applied as foliar spray or through drip-irrigation lines, on bacterial wilt incidence and yield of grafted tomatoes. The experiments were conducted in a field with race 1, biovar 1 strain of Ralstonia solanacearum, causal agent of tomato bacterial wilt. In all three experiments, the susceptible tomato variety BHN 602, grafted onto a resistant rootstock BHN 998, was compared with nongrafted BHN 602, treated with or without foliar applications of ASM and with grafted plants treated with foliar applications of ASM. In two experiments, an additional treatment of drip applications of ASM on grafted and nongrafted plants was evaluated. Grafting alone or in combination with drip applications of ASM (178.6 μM) significantly reduced disease incidence and increased total marketable yield relative to nongrafted treatments. There were no significant differences between grafted plants with or without drip ASM applications in terms of bacterial wilt incidence or total marketable yield. However, we demonstrate for the first time that foliar ASM applications on grafted plants negatively affects the total marketable yield compared with drip ASM applications on grafted plants or nontreated grafted control.

Inducible expression of Bs2 R gene from Capsicum chacoense in sweet orange (Citrus sinensis L. Osbeck) confers enhanced resistance to citrus canker disease.

Transgenic expression of the pepper Bs2 gene confers resistance to Xanthomonas campestris pv. vesicatoria (Xcv) pathogenic strains which contain the avrBs2 avirulence gene in susceptible pepper and tomato varieties. The avrBs2 gene is highly conserved among members of the Xanthomonas genus, and the avrBs2 of Xcv shares 96% homology with the avrBs2 of Xanthomonas citri subsp. citri (Xcc), the causal agent of citrus canker disease. A previous study showed that the transient expression of pepper Bs2 in lemon leaves reduced canker formation and induced plant defence mechanisms. In this work, the effect of the stable expression of Bs2 gene on citrus canker resistance was evaluated in transgenic plants of Citrus sinensis cv. Pineapple. Interestingly, Agrobacterium-mediated transformation of epicotyls was unsuccessful when a constitutive promoter (2× CaMV 35S) was used in the plasmid construction, but seven transgenic lines were obtained with a genetic construction harbouring Bs2 under the control of a pathogen-inducible promoter, from glutathione S-transferase gene from potato. A reduction of disease symptoms of up to 70% was observed in transgenic lines expressing Bs2 with respect to non-transformed control plants. This reduction was directly dependent on the Xcc avrBs2 gene since no effect was observed when a mutant strain of Xcc with a disruption in avrBs2 gene was used for inoculations. Additionally, a canker symptom reduction was correlated with levels of the Bs2 expression in transgenic plants, as assessed by real-time qPCR, and accompanied by the production of reactive oxygen species. These results indicate that the pepper Bs2 resistance gene is also functional in a family other than the Solanaceae, and could be considered for canker control.