Allelic variations in the chpG effector gene within Clavibacter michiganensis populations determine pathogen host range.

Plant pathogenic bacteria often have a narrow host range, which can vary among different isolates within a population. Here, we investigated the host range of the tomato pathogen Clavibacter michiganensis (Cm). We determined the genome sequences of 40 tomato Cm isolates and screened them for pathogenicity on tomato and eggplant. Our screen revealed that out of the tested isolates, five were unable to cause disease on any of the hosts, 33 were exclusively pathogenic on tomato, and two were capable of infecting both tomato and eggplant. Through comparative genomic analyses, we identified that the five non-pathogenic isolates lacked the chp/tomA pathogenicity island, which has previously been associated with virulence in tomato. In addition, we found that the two eggplant-pathogenic isolates encode a unique allelic variant of the putative serine hydrolase chpG (chpGC), an effector that is recognized in eggplant. Introduction of chpGC into a chpG inactivation mutant in the eggplant-non-pathogenic strain Cm101, failed to complement the mutant, which retained its ability to cause disease in eggplant and failed to elicit hypersensitive response (HR). Conversely, introduction of the chpG variant from Cm101 into an eggplant pathogenic Cm isolate (C48), eliminated its pathogenicity on eggplant, and enabled C48 to elicit HR. Our study demonstrates that allelic variation in the chpG effector gene is a key determinant of host range plasticity within Cm populations.

New insight into protective effect against oxidative stress and biosynthesis of exopolysaccharides produced by Lacticaseibacillus paracasei NC4 from fermented eggplant.

Fermented eggplant is a traditional fermented food, however lactic acid bacteria capable of producing exopolysaccharide (EPS) have not yet been exploited. The present study focused on the production and protective effects against oxidative stress of an EPS produced by Lacticaseibacillus paracasei NC4 (NC4-EPS), in addition to deciphering its genomic features and EPS biosynthesis pathway. Among 54 isolates tested, strain NC4 showed the highest EPS yield and antioxidant activity. The maximum EPS production (2.04 ± 0.11 g/L) was achieved by culturing in MRS medium containing 60 g/L sucrose at 37 °C for 48 h. Under 2 mM H2O2 stress, the survival of a yeast model Saccharomyces cerevisiae treated with 0.4 mg/mL NC4-EPS was 2.4-fold better than non-treated cells, which was in agreement with the catalase and superoxide dismutase activities measured from cell lysates. The complete genome of NC4 composed of a circular chromosome of 2,888,896 bp and 3 circular plasmids. The NC4 genome comprises more genes with annotated function in nitrogen metabolism, phosphorus metabolism, cell division and cell cycle, and iron acquisition and metabolism as compared to other reported L. paracasei. Of note, the eps gene cluster is not conserved across L. paracasei. Pathways of sugar metabolism for EPS biosynthesis were proposed for the first time, in which gdp pathway only present in few plant-derived bacteria was identified. These findings shed new light on the cell-protective activity and biosynthesis of EPS produced by L. paracasei, paving the way for future efforts to enhance yield and tailor-made EPS production for food and pharmaceutical industries.

Comprehensive analysis of little leaf disease incidence and resistance in eggplant.

BACKGROUND: Little leaf disease caused by phytoplasma infection is a significant threat to eggplant (also known as brinjal) cultivation in India. This study focused on the molecular characterisation of the phytoplasma strains and insect vectors responsible for its transmission and screening of brinjal germplasm for resistance to little leaf disease. RESULTS: Surveys conducted across districts in the Tamil Nadu state of India during 2021-2022 showed a higher incidence of phytoplasma during the Zaid (March to June), followed by Kharif (June to November) and Rabi (November to March) seasons with mean incidence ranging from 22 to 27%. As the name indicates, phytoplasma infection results in little leaf (reduction in leaf size), excessive growth of axillary shoots, virescence, phyllody, stunted growth, leaf chlorosis and witches' broom symptoms. PCR amplification with phytoplasma-specific primers confirmed the presence of this pathogen in all symptomatic brinjal plants and in Hishimonus phycitis (leafhopper), providing valuable insights into the role of leafhoppers in disease transmission. BLAST search and phylogenetic analysis revealed the phytoplasma strain as "Candidatus Phytoplasma trifolii". Insect population and disease dynamics are highly influenced by environmental factors such as temperature, relative humidity and rainfall. Further, the evaluation of 22 eggplant accessions revealed immune to highly susceptible responses where over 50% of the entries were highly susceptible. Finally, additive main effect and multiplicative interaction (AMMI) and won-where biplot analyses identified G18 as a best-performing accession for little leaf resistance due to its consistent responses across multiple environments. CONCLUSIONS: This research contributes essential information on little leaf incidence, symptoms, transmission and resistance profiles of different brinjal genotypes, which together ensure effective and sustainable management of this important disease of eggplants.

Map-based cloning of LPD, a major gene positively regulates leaf prickle development in eggplant.

KEY MESSAGE: A critical gene for leaf prickle development (LPD) in eggplant was mapped on chromosome E06 and was confirmed to be SmARF10B through RNA interference using a new genetic transformation technique called SACI developed in this study Prickles on eggplant pose challenges for agriculture and are undesirable in cultivated varieties. This study aimed to uncover the genetic mechanisms behind prickle formation in eggplant. Using the F2 and F2:3 populations derived from a cross between the prickly wild eggplant, YQ, and the prickle-free cultivated variety, YZQ, we identified a key genetic locus (LPD, leaf prickle development) on chromosome E06 associated with leaf prickle development through BSA-seq and QTL mapping. An auxin response factor gene, SmARF10B, was predicted as the candidate gene as it exhibited high expression in YQ's mature leaves, while being significantly low in YZQ. Downregulating SmARF10B in YQ through RNAi using a simple and efficient Agrobacterium-mediated genetic transformation method named Seedling Apical Cut Infection (SACI) developed in this study substantially reduced the size and density of leaf prickles, confirming the role of this gene in prickle development. Besides, an effective SNP was identified in SmARF10B, resulting in an amino acid change between YQ and YZQ. However, this SNP did not consistently correlate with prickle formation in eight other eggplant materials examined. This study sheds light on the pivotal role of SmARF10B in eggplant prickle development and introduces a new genetic transformation method for eggplant, paving the way for future research in this field.

Aquincola agrisoli sp. nov., isolated from rhizospheric soil of eggplant and in silico genome mining for the prediction of biosynthetic gene clusters.

A Gram-stain negative, aerobic, rod-shaped, motile and flagellated novel bacterial strain, designated MAHUQ-54T, was isolated from the rhizospheric soil of eggplant. The colonies were observed to be light pink coloured, smooth, spherical and 0.2-0.6 mm in diameter when grown on R2A agar medium for 2 days. MAHUQ-54T was able to grow at 15-40 °C, at pH 5.5-9.0 and in the presence of 0-0.5 % NaCl (w/v). The strain gave positive results for both catalase and oxidase tests. The strain was positive for hydrolysis of l-tyrosine, urea, Tween 20 and Tween 80. On the basis of the results of 16S rRNA gene sequence comparisons, the isolate was identified as a member of the genus Aquincola and is closely related to Aquincola tertiaricarbonis L10T (98.8 % sequence similarity) and Leptothrix mobilis Feox-1T (98.2 %). MAHUQ-54T has a draft genome size of 5 994 516 bp (60 contigs), annotated with 5348 protein-coding genes, 45 tRNA and 5 rRNA genes. The average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) values between MAHUQ-54T and its closest phylogenetic neighbours were 75.8-83.3 and 20.8-25.3 %, respectively. In silico genome mining revealed that MAHUQ-54T has a significant potential for the production of novel natural products in the future. The genomic DNA G+C content was determined to be 70.4 %. The predominant isoprenoid quinone was ubiquinone-8. The major fatty acids were identified as C16  :  0, summed feature 3 (comprising C16  :  1ω7c and/or C16  :  1ω6c) and summed feature 8 (comprising C18  :  1ω7c and/or C18  :  1ω6c). On the basis of dDDH, ANI value, genotypic analysis, chemotaxonomic and physiological data, strain MAHUQ-54T represents a novel species within the genus Aquincola, for which the name Aquincola agrisoli sp. nov. is proposed, with MAHUQ-54T (=KACC 22001T = CGMCC 1.18515T) as the type strain.

Potential of biosynthesized zinc oxide nanoparticles to control Fusarium wilt disease in eggplant (Solanum melongena) and promote plant growth.

In this study, a novel, non-toxic, eco-friendly zinc oxide nanoparticles (ZnO-NPs) was used instead of the synthetic fungicides widely used to control the destructive phytopathogenic fungus Fusarium oxysporum, the causative agent of wilt disease in Solanum melongena L. Herein, the biosynthesized ZnO-NPs was carried out by Penicillium expansum ATCC 7861. In vitro, mycosynthesized ZnO-NPs exhibited antifungal activity against Fusarium oxysporum. In vivo, ZnO-NPs suppressed Fusarium wilt disease in cultivated Solanum melongena L. by decreasing the disease severity with 75% of plant protection. Moreover, ZnO-NPs stimulated the recovery of eggplant as an indicated by improving of morphological and metabolic indicators including plant height(152.5%), root length(106.6%), plant fresh biomass (146%), chlorophyll a (102.8%), chlorophyll b (67.86%), total soluble carbohydrates (48.5%), total soluble protein (81.8%), phenol (10.5%), antioxidant activity and isozymes compared with infected control. Therefore, this study suggests using mycosynthesized ZnO-NPs as an alternative to synthetic fungicides not only to eradicate the Fusarium wilt disease in cultivated eggplant (Solanum melongena) but also to promote the growth parameters and metabolic aspects.

Biocontrol Streptomyces Induces Resistance to Bacterial Wilt by Increasing Defense-Related Enzyme Activity in Solanum melongena L.

Streptomyces strains were isolated from rhizosphere soil and evaluated for in vitro plant growth and antagonistic potential against Ralstonia solanacearum. Based on their in vitro screening, seven Streptomyces were evaluated for plant growth promotion (PGP) and biocontrol efficacy by in-planta and pot culture study. In the in-planta study, Streptomyces-treated eggplant seeds showed better germination percentage, plant growth, and disease occurrence against R. solanacearum than the control treatment. Hence, all seven Streptomyces cultures were developed as a bioformulation by farmyard manure and used for pot culture study. The highest plant growth, weight, and total chlorophyll content were observed in UP1A-1-treated eggplant followed by UP1A-4, UT4A-49, and UT6A-57. Similarly, the maximum biocontrol efficacy was observed in UP1A-1-treated eggplants against bacterial wilt. The biocontrol potential of Streptomyces is also confirmed through metabolic responses by assessing the activities of the defense-related enzymes peroxidase (POX), polyphenol oxidase (PPO), and phenylalanine ammonia-lyase (PAL) and as well as the levels of total phenol. Treatment with UP1A-1/ UT4A-49 and challenge with R. solanacearum led to maximum changes in the activities of POX, PPO, and PAL and the levels of total phenol in the eggplants at different time intervals. Alterations in enzymes of UP1A-1 treatment were related to early defense responses in eggplant. Therefore, the treatment with UP1A-1 significantly delayed the establishment of bacterial wilt in eggplant. Altogether, the present study suggested that the treatment of Streptomyces maritimus UP1A-1 fortified farmyard manure has improved the plant growth and stronger disease control against R. solanacearum on eggplant.

Structural insights into the lipid transfer mechanism of a non-specific lipid transfer protein.

The non-specific lipid transfer proteins (nsLTPs) are multifunctional seed proteins engaged in several different physiological processes. The nsLTPs are stabilized by four disulfide bonds and exhibit a characteristic hydrophobic cavity, which is the primary lipid binding site. While these proteins are known to transfer lipids between membranes, the mechanism of lipid transfer has remained elusive. Four crystal structures of nsLTP from Solanum melongena, one in the apo-state and three myristic acid bound states were determined. Among the three lipid bound states, two lipid molecules were bound on the nsLTP surface at different positions and one was inside the cavity. The lipid-dependent conformational changes leading to opening of the cavity were revealed based on structural and spectroscopic data. The surface-bound lipid represented a transient intermediate state and the lipid ultimately moved inside the cavity through the cavity gate as revealed by molecular dynamics simulations. Two critical residues in the loop regions played possible 'gating' role in the opening and closing of the cavity. Antifungal activity and membrane permeabilization effect of nsLTP against Fusarium oxysporum suggested that it could possibly involve in bleaching out the lipids. Collectively, these studies support a model of lipid transfer mechanism by nsLTP via intermediate states.

Weighted Gene Co-Expression Analysis Network-Based Analysis on the Candidate Pathways and Hub Genes in Eggplant Bacterial Wilt-Resistance: A Plant Research Study.

Solanum melongena L. (eggplant) bacterial wilt is a severe soil borne disease. Here, this study aimed to explore the regulation mechanism of eggplant bacterial wilt-resistance by transcriptomics with weighted gene co-expression analysis network (WGCNA). The different expression genes (DEGs) of roots and stems were divided into 21 modules. The module of interest (root: indianred4, stem: coral3) with the highest correlation with the target traits was selected to elucidate resistance genes and pathways. The selected module of roots and stems co-enriched the pathways of MAPK signalling pathway, plant pathogen interaction, and glutathione metabolism. Each top 30 hub genes of the roots and stems co-enriched a large number of receptor kinase genes. A total of 14 interesting resistance-related genes were selected and verified with quantitative polymerase chain reaction (qPCR). The qPCR results were consistent with those of WGCNA. The hub gene of EGP00814 (namely SmRPP13L4) was further functionally verified; SmRPP13L4 positively regulated the resistance of eggplant to bacterial wilt by qPCR and virus-induced gene silencing (VIGS). Our study provides a reference for the interaction between eggplants and bacterial wilt and the breeding of broad-spectrum and specific eggplant varieties that are bacterial wilt-resistant.

Combining genome composition and differential gene expression analyses reveals that SmPGH1 contributes to bacterial wilt resistance in somatic hybrids.

KEY MESSAGE: Clarification of the genome composition of the potato + eggplant somatic hybrids cooperated with transcriptome analysis efficiently identified the eggplant gene SmPGH1 that contributes to bacterial wilt resistance. The cultivated potato is susceptible and lacks resistance to bacterial wilt (BW), a soil-borne disease caused by Ralstonia solanacearum. It also has interspecies incompatibility within Solanaceae plants. Previously, we have successfully conducted the protoplast fusion of potato and eggplant and regenerated somatic hybrids that showing resistance to eggplant BW. For efficient use of these novel germplasm and improve BW resistance of cultivated potato, it is essential to dissect the genetic basis of the resistance to BW obtained from eggplant. The strategy of combining genome composition and transcriptome analysis was established to explore the gene that confers BW resistance to the hybrids. Genome composition of the 90 somatic hybrids was studied using genomic in situ hybridization coupled with 44 selected eggplant-specific SSRs (smSSRs). The analysis revealed a diverse set of genome combinations among the hybrids and showed a possibility of integration of alien genes along with the detection of 7 smSSRs linked to BW resistance (BW-linked SSRs) in the hybrids. Transcriptome comparison between the resistant and susceptible gene pools identified a BW resistance associated gene, smPGH1, which was significantly induced by R. solanacearum in the resistant pool. Remarkably, smPGH1 was co-localized with the BW-linked SSR emh01E15 on eggplant chromosome 9, which was further confirmed that smPGH1 was activated by R. solanacearum only in the resistant hybrids. Taken together, the identified gene smPGH1 and BW-linked SSRs have provided novel genetic resources that will aid in potato breeding for BW resistance.

Engineered gray mold resistance, antioxidant capacity, and pigmentation in betalain-producing crops and ornamentals.

Betalains are tyrosine-derived red-violet and yellow plant pigments known for their antioxidant activity, health-promoting properties, and wide use as food colorants and dietary supplements. By coexpressing three genes of the recently elucidated betalain biosynthetic pathway, we demonstrate the heterologous production of these pigments in a variety of plants, including three major food crops: tomato, potato, and eggplant, and the economically important ornamental petunia. Combinatorial expression of betalain-related genes also allowed the engineering of tobacco plants and cell cultures to produce a palette of unique colors. Furthermore, betalain-producing tobacco plants exhibited significantly increased resistance toward gray mold (Botrytis cinerea), a pathogen responsible for major losses in agricultural produce. Heterologous production of betalains is thus anticipated to enable biofortification of essential foods, development of new ornamental varieties, and innovative sources for commercial betalain production, as well as utilization of these pigments in crop protection.

The eggplant transcription factor MYB44 enhances resistance to bacterial wilt by activating the expression of spermidine synthase.

Bacterial wilt (BW) caused by Ralstonia solanacearum is a serious disease affecting the production of Solanaceae species, including eggplant (Solanum melongena). However, few resistance genes have been identified in eggplant, and therefore the underlying mechanism of BW resistance remains unclear. Hence, we investigated a spermidine synthase (SPDS) gene from eggplant and created knock-down lines with virus-induced gene silencing. After eggplant was infected with R. solanacearum, the SmSPDS gene was induced, concurrent with increased spermidine (Spd) content, especially in the resistant line. We speculated that Spd plays a significant role in the defense response of eggplant to BW. Moreover, using the yeast one-hybrid approach and dual luciferase-based transactivation assay, an R2R3-MYB transcription factor, SmMYB44, was identified as directly binding to the SmSPDS promoter, activating its expression. Overexpression of SmMYB44 in eggplant induced the expression of SmSPDS and Spd content, increasing the resistance to BW. In contrast, the SmMYB44-RNAi transgenic plants showed more susceptibility to BW compared with the control plants. Our results provide insight into the SmMYB44-SmSPDS-Spd module involved in the regulation of resistance to R. solanacearum. This research also provides candidates to enhance resistance to BW in eggplant.

Specific PCR-based detection of Phomopsis vexans the cause of leaf blight and fruit rot pathogen of Solanum melongena L.

Leaf blight and fruit rot disease caused by Phomopsis vexans is a devastating disease of brinjal. The detection of P. vexans in plant parts and seeds of brinjal can be complicated, mainly when the inoculum is present at low levels and/or overgrown by fast-growing saprophytic fungi or other seed-borne fungi. A PCR-based diagnostic method was developed with specific primers designed based on sequence data of a region consisting of the 5·8S RNA gene and internal transcribed spacers, ITS 1 and ITS 2 of nuclear ribosomal RNA gene (rDNA) repeats of P. vexans. The efficiency and specificity of primer pairs PvexF/PvexR designed were established by PCR analysis of DNA from P. vexans strains isolated from India and fungal isolates of other genera. A single amplification product of 323-bp was detected from DNA of P. vexans isolates. No cross-reaction was observed with any of the other isolates tested. The specific primers designed and employed in PCR detected P. vexans up to 10 pg from DNA isolated from pure culture. This is the first report on the development of species-specific PCR assay for identification and detection of P. vexans. Thus, PCR-based assay developed is very specific, rapid, confirmatory and sensitive tool for the detection of pathogen P. vexans at early stages. SIGNIFICANCE AND IMPACT OF THE STUDY: Phomopsis vexans is an important seed-borne pathogenic fungus responsible for leaf blight and fruit rot in brinjal. Current detection methods, based on culture and morphological identification is time consuming, laborious and are not always reliable. A PCR-based diagnostic method was developed with species-specific primers designed based on sequence data of a region consisting of the 5·8S RNA gene and internal transcribed spacers, ITS 1 and ITS 2 of nuclear ribosomal RNA gene (rDNA) repeats of P. vexans.

Antigenic and pathogenicity activities of Ralstonia solanacearum race 3 biovar 2 molecularly identified and detected by indirect ELISA using polyclonal antibodies generated in rabbits.

Eight molecular-characterized isolates of Ralstonia solanacearum from potato belonging to race 3 biovar 2, their virulence were evaluated on potato cv. Lady Rosette, tomato cv. Strain B, eggplant cv. Balady and pepper cv. Balady and showed high virulence on potato and tomato, and lower virulence on eggplant and pepper. A laboratory study conducted to produce polyclonal antibodies against the potato brown rot bacterium; R. solanacearum cells were generated in female New Zealand white rabbits. A modification were made on the technique of indirect enzyme-linked immunosorbent assay (ELISA) to improve the sensitivity of detection, including antigenic and sensitivity to R. solanacearum race 3 biovar 2 isolates. Determination of the optimum period to collect the antiserum (including, polyclonal antibodies) showed that the best collection dates were at 14, 3 and 7 days, in that order. The efficiency of the antiserum was compared among 42 isolates that cause potato brown rot disease; our polyclonal antiserum (14 days) reacted positively with all tested isolates at a dilution of 1:6.4 × 103. Data indicated the different reactions of eight R. solanacearum isolates at various dilutions (1:1.6 × 103 to 1:5.12 × 106) at 14 days against polyclonal antiserumat a concentration of approximately 1 × 108 CFU/mL and we found the lowest detection level by the indirect ELISA technique was 106 CFU/mL. Finally we recommended the reasonable sensitivity results of the ELISA technique to detect the bacterial pathogen given than the cost of this technique if much lower than that of other expensive molecular techniques.

Characterization and Synergistic Effect of Antifungal Volatile Organic Compounds Emitted by the Geotrichum candidum PF005, an Endophytic Fungus from the Eggplant.

Plant-associated endophytes are recognized as sources of novel bioactive molecules having diverse applications. In this study, an endophytic yeast-like fungal strain was isolated from the fruit of eggplant (Solanum melongena) and identified as Geotrichum candidum through phenotypic and genotypic characterizations. This endophytic G. candidum isolate PF005 was found to emit fruity scented volatiles. The compositional profiling of volatile organic compounds (VOCs) revealed the presence of 3-methyl-1-butanol, ethyl 3-methylbutanoate, 2-phenylethanol, isopentyl acetate, naphthalene, and isobutyl acetate in significant proportion when analyzed on a time-course basis. The VOCs from G. candidum exhibited significant mycelial growth inhibition (54%) of phytopathogen Rhizoctonia solani, besides having mild antifungal activity against a few other fungi. The source of carbon as a nutrient was found to be an important factor for the enhanced biosynthesis of antifungal VOCs. The antifungal activity against phytopathogen R. solani was improved up to 91% by feeding the G. candidum with selective precursors of alcohol and ester volatiles. Furthermore, the antifungal activity of VOCs was enhanced synergistically up to 92% upon the exogenous addition of naphthalene (1.0 mg/plate). This is the first report of G. candidum as an endophyte emitting antifungal VOCs, wherein 2-penylethanol, isopentyl acetate, and naphthalene were identified as important contributors to its antifungal activity. Possible utilization of G. candidum PF005 as a mycofumigant has been discussed based upon its antifungal activity and the qualified presumption of safety status.

Host range and molecular characterization of a lytic Pradovirus-like Ralstonia phage RsoP1IDN isolated from Indonesia.

A lytic Ralstonia solanacearum-infecting phage designated Ralstonia phage RsoP1IDN was isolated from soil in Indonesia. The phage has a linear double-stranded DNA genome of 41,135 bp with 413-bp terminal repeats, and contains 41 annotated open reading frames. The phage is most closely related to Ralstonia phage RSB1, but different from RSB1 mainly in containing a putative HNH homing endonuclease and having a narrower host range. Our phylogenetic and genomic analyses revealed that both phages RsoP1IDN and RSB1 belong to the genus Pradovirus or a new genus, and not Phikmvvirus as previously reported for phage RSB1. RsoP1IDN is the first sequenced and characterized R. solanacearum-infecting phage isolated from Indonesia in the proposed species Ralstonia virus RsoP1IDN.

Soil Microbiomes Associated with Verticillium Wilt-Suppressive Broccoli and Chitin Amendments are Enriched with Potential Biocontrol Agents.

Two naturally infested Verticillium wilt-conducive soils from the Salinas Valley of coastal California were amended with disease-suppressive broccoli residue or crab meal amendments, and changes to the soil prokaryote community were monitored using Illumina sequencing of a 16S ribosomal RNA gene library generated from 160 bulk soil samples. The experiment was run in a greenhouse, twice, with eggplant as the Verticillium wilt-susceptible host. Disease suppression, plant height, soil microsclerotia density, and soil chitinase activity were assessed at the conclusion of each experiment. In soil with high microsclerotia density, all amendments significantly reduced Verticillium wilt severity and microsclerotia density, and increased soil chitinase activity. Plant height was increased only in the broccoli-containing treatments. In total, 8,790 error-corrected sequence variants representing 1,917,893 different sequences were included in the analyses. The treatments had a significant impact on the soil microbiome community structure but measures of α diversity did not vary between treatments. Community structure correlated with disease score, plant height, microsclerotia density, and soil chitinase activity, suggesting that the prokaryote community may affect the disease-related response variables or vice versa. Similarly, the abundance of 107 sequence variants correlated with disease-related response variables, which included variants from genera with known antagonists of filamentous fungal plant pathogens, such as Pseudomonas and Streptomyces. Overall, genera with antifungal antagonists were more abundant in amended soils than unamended soils, and constituted up to 8.9% of all sequences in broccoli+crabmeal-amended soil. This study demonstrates that substrate-mediated shifts in soil prokaryote communities are associated with the transition of Verticillium wilt-conducive soils to Verticillium wilt-suppressive soils, and suggests that soils likely harbor numerous additional antagonists of fungal plant pathogens that contribute to the biological suppression of plant disease.

Epigenetic regulation of the expression of WRKY75 transcription factor in response to biotic and abiotic stresses in Solanaceae plants.

KEY MESSAGE: SlyWRKY75: gene expression was induced in response to biotic stresses, especially in Botrytis cinerea-infected tomato plants, in which Sly-miR1127-3p is a putative SlyWRKY75 regulator and epigenetic marks were detected. WRKY75 transcription factor involved in Pi homeostasis was recently found also induced in defense against necrotrophic pathogens. In this study, we analyzed by RT-qPCR the expression of SlyWRKY75 gene in tomato plants in response to abiotic stresses (drought or heat) and biotic stresses (Colorado potato beetle larvae infestation, Pseudomonas syringae or Botrytis cinerea infection) being only differentially expressed following biotic stresses, especially upon B. cinerea infection (55-fold induction). JA and JA-Ile levels were significantly increased in tomato plants under biotic stresses compared with control plants, indicating that SlyWRKY75 might be a transcriptional regulator of the JA pathway. The contribution of miRNAs and epigenetic molecular mechanisms to the regulation of this gene in B. cinerea-infected tomato plants was explored. We identified a putative Sly-miR1127-3p miRNA predicted to bind the intronic region of the SlyWRKY75 genomic sequence. Sly-miR1127-3p miRNA was repressed in infected plants (0.4-fold) supporting that it might act as an epigenetic regulation factor of SlyWRKY75 gene expression rather than via the post-transcriptional mechanisms of canonical miRNAs. It has been proposed that certain miRNAs can mediate DNA methylation in the plant nucleus broadening miRNA functions with transcriptional gene silencing by targeting intron-containing pre-mRNAs. Histone modifications analysis by chromatin immunoprecipitation (ChIP) demonstrated the presence of the activator histone modification H3K4me3 on SlyWRKY75 transcription start site and gene body. The induction of this gene in response to B. cinerea correlates with the presence of an activator mark. Thus, miRNAs and chromatin modifications might cooperate as epigenetic factors to modulate SlyWRKY75 gene expression.

Assessment of edibility and effect of arbuscular mycorrhizal fungi on Solanum melongena L. grown under heavy metal(loid) contaminated soil.

Arbuscular mycorrhizal fungi (AMF) aids in plant establishment at heavy metal(loid) (HM) contaminated soils, strengthening plant defense system along with promoting growth. A pot experiment was carried out to evaluate the effect of AMF on eggplants grown under HM stress. Further, the potential health risks of HM exposure to the humans via dietary intake of eggplant were also estimated. Results showed that AMF application improved growth, biomass and antioxidative defense response of plants against HM stress. Significant difference (p ≤ 0.001) in parameters under study was found on increasing metal dose and on application of AMF. Among metal(loid)s maximum uptake was recorded for Pb (29.64mgkg-1 in roots; 23.08mgkg-1 in shoot) followed by As (3.84mgkg-1 in roots; 8.20mgkg-1 in shoot) and, Cd (0.96mgkg-1 in roots; 2.12mgkg-1 in shoot). Based on the accumulation of HM in edible part, Hazard Quotient (HQ) was calculated. HQ was found to be > 1 for Pb, which highlights the risks associated with consumption of Eggplants grown on Pb contaminated soil. However this potential, which was further enhanced by application of AMF, can be harnessed for on-site remediation of Pb contaminated soils. The content of Cd and As in the edible part was found to be within safe limits (HQ < 1) when compared to chronic reference dose stated by USEPA.

Phylogenetic Assignment of Ralstonia pseudosolanacearum (Ralstonia solanacearum Phylotype I) Isolated from Rosa spp.

During the last two years, greenhouse cultivation of rose (Rosa spp.) in the Netherlands has been challenged by an uncommon bacterial disease. Affected plants suffered from chlorosis, stunting, wilting, and necrosis. The bacterial isolates obtained from the different Rosa spp. cultivars were all identified as phylotype I, sequevar 33 from the 'Ralstonia solanacearum species complex' (RSSC), actually reclassified as Ralstonia pseudosolanacearum. The work in this paper considers the genetic diversity and the phylogenetic position of 129 R. pseudosolanacearum isolates from the outbreak. This was assessed by AFLP based on four different primer combinations and MLP using partial sequences of the egl, mutS, and fliC genes. The AFLP revealed identical profiles for all the isolates, irrespective of their association with Rosa sp. propagating material, Rosa spp. plants for cut flowers, or water used in the different greenhouse cultivations. These AFLP profiles were unique and diverged from profiles of all other reference isolates in the RSSC included. Furthermore, MLP on egl, fliC, and mutS gene sequences clearly demonstrated that all R. pseudosolanacearum isolates clustered in phylotype I, as a distinct monophyletic group. Interestingly, this monophyletic group also included phylotype I strain Rs-09-161 from eggplant (Solanum melongena), isolated in 2009 in India. AFLP and MLP were both efficient in revealing the genetic divergence from the RSSC isolates included. The phylogenetic tree constructed from the AFLP profiles was, in general, in agreement with the one obtained from MLP. Both phylogenetic trees displayed a similar clustering, supported by high posterior probabilities. Both methodologies clearly demonstrated that the R. pseudosolanacearum isolates from Rosa spp. grouped in a monophyletic group inside phylotype I, with a particular correspondence to a strain present in India, as revealed in MLP.