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Understanding the response of soil microbial communities to pathogenic Ralstonia solanacearum is crucial for preventing bacterial wilt outbreaks. In this study, we investigated the soil physicochemical and microbial community to assess their impact on the pathogenic R.solanacearum through metagenomics. Our results revealed that certain archaeal taxa were the main contributors influencing the health of plants. Additionally, the presence of the pathogen showed a strong negative correlation with soil phosphorus levels, while soil phosphorus was significantly correlated with bacterial and archaeal communities. We found that the network of microbial interactions in healthy plant rhizosphere soils was more complex compared to diseased soils. The diseased soil network had more linkages, particularly related to the pathogen occurrence. Within the network, the family Comamonadaceae, specifically Ramlibacter_tataouinensis, was enriched in healthy samples and showed a significantly negative correlation with the pathogen. In terms of archaea, Halorubrum, Halorussus_halophilus (family: Halobacteriaceae), and Natronomonas_pharaonis (family: Haloarculaceae) were enriched in healthy plant rhizosphere soils and showed negative correlations with R.solanacearum. These findings suggested that the presence of these archaea may potentially reduce the occurrence of bacterial wilt disease. On the other hand, Halostagnicola_larseniia and Haloterrigena_sp._BND6 (family: Natrialbaceae) had higher relative abundance in diseased plants and exhibited significantly positive correlations with R.solanacearum, indicating their potential contribution to the pathogen's occurrence. Moreover, we explored the possibility of functional gene sharing among the correlating bacterial pairs within the Molecular Ecological Network. Our analysis revealed 468 entries of horizontal gene transfer (HGT) events, emphasizing the significance of HGT in shaping the adaptive traits of plant-associated bacteria, particularly in relation to host colonization and pathogenicity. Overall, this work revealed key factors, patterns and response mechanisms underlying the rhizosphere soil microbial populations. The findings offer valuable guidance for effectively controlling soil-borne bacterial diseases and developing sustainable agriculture practices. [ABSTRACT FROM AUTHOR]

CRISPR/Cas is a breakthrough genome editing system because of its precision, target specificity, and efficiency. As a speed breeding system, it is more robust than the conventional breeding and biotechnological approaches for qualitative and quantitative trait improvement. Tomato (Solanum lycopersicum L.) is an economically important crop, but its yield and productivity have been severely impacted due to different abiotic and biotic stresses. The recently identified SlHyPRP1 and SlDEA1 are two potential negative regulatory genes in response to different abiotic (drought and salinity) and biotic stress (bacterial leaf spot and bacterial wilt) conditions in S. lycopersicum L. The present study aimed to evaluate the drought, salinity, bacterial leaf spot, and bacterial wilt tolerance response in S. lycopersicum L. crop through CRISPR/Cas9 genome editing of SlHyPRP1 and SlDEA1 and their functional analysis. The transient single- and dual-gene SlHyPRP1 and SlDEA1 CRISPR-edited plants were phenotypically better responsive to multiple stress factors taken under the study. The CRISPRedited SlHyPRP1 and SlDEA1 plants showed a higher level of chlorophyll and proline content compared to wild-type (WT) plants under abiotic stress conditions. Reactive oxygen species accumulation and the cell death count per total area of leaves and roots under biotic stress were less in CRISPR-edited SlHyPRP1 and SlDEA1 plants compared to WT plants. The study reveals that the combined loss-of-function of SlHyPRP1 along with SlDEA1 is essential for imparting significant multi-stress tolerance (drought, salinity, bacterial leaf spot, and bacterial wilt) in S. lycopersicum L. The main feature of the study is the detailed genetic characterization of SlDEA1, a poorly studied 8CM family gene in multi-stress tolerance, through the CRISPR/Cas9 gene editing system. The study revealed the key negative regulatory role of SlDEA1 that function together as an anchor gene with SlHyPRP1 in imparting multi-stress tolerance in S. lycopersicum L. It was interesting that the present study also showed that transient CRISPR/Cas9 editing events of SlHyPRP1 and SlDEA1 genes were successfully replicated in stably generated parent-genome-edited line (GEd0) and genome-edited first-generation lines (GEd1) of S. lycopersicum L. With these upshots, the study's key findings demonstrate outstanding value in developing sustainable multi-stress tolerance in S. lycopersicum L. and other crops to cope with climate change. [ABSTRACT FROM AUTHOR]

The soil microbial community plays a critical role in promoting robust plant growth and serves as an effective defence mechanism against root pathogens. Current research has focused on unravelling the compositions and functions of diverse microbial taxa in plant rhizospheres invaded by Ralstonia solanacearum, however, the specific mechanisms by which key microbial groups with distinct functions exert their effects remain unclear. In this study, we employed a combination of amplicon sequencing and metabolomics analysis to investigate the principal metabolic mechanisms of key microbial taxa in plant rhizosphere soil. Compared to the healthy tobacco rhizosphere samples, the bacterial diversity and co-occurrence network of the diseased tobacco rhizosphere soil were significantly reduced. Notably, certain genera, including Gaiella, Rhodoplanes, and MND1 (Nitrosomonadaceae), were found to be significantly more abundant in the rhizosphere of healthy plants than in that of diseased plants. Eight environmental factors, including exchangeable magnesium, available phosphorus, and pH, were found to be crucial factors influencing the composition of the microbial community. Ralstonia displayed negative correlations with pH, exchangeable magnesium, and cation exchange flux, but showed a positive correlation with available iron. Furthermore, metabolomic analysis revealed that the metabolic pathways related to the synthesis of various antibacterial compounds were significantly enriched in the healthy group. The correlation analysis results indicate that the bacterial genera Polycyclovorans, Lysobacter, Pseudomonas, and Nitrosospira may participate in the synthesis of antibacterial compounds. Collectively, our findings contribute to a more in-depth understanding of disease resistance mechanisms within healthy microbial communities and provide a theoretical foundation for the development of targeted strategies using beneficial microorganisms to suppress disease occurrence. [ABSTRACT FROM AUTHOR]

CRISPR/Cas is a breakthrough genome editing system because of its precision, target specificity, and efficiency. As a speed breeding system, it is more robust than the conventional breeding and biotechnological approaches for qualitative and quantitative trait improvement. Tomato (Solanum lycopersicum L.) is an economically important crop, but its yield and productivity have been severely impacted due to different abiotic and biotic stresses. The recently identified SlHyPRP1 and SlDEA1 are two potential negative regulatory genes in response to different abiotic (drought and salinity) and biotic stress (bacterial leaf spot and bacterial wilt) conditions in S. lycopersicum L. The present study aimed to evaluate the drought, salinity, bacterial leaf spot, and bacterial wilt tolerance response in S. lycopersicum L. crop through CRISPR/Cas9 genome editing of SlHyPRP1 and SlDEA1 and their functional analysis. The transient single- and dual-gene SlHyPRP1 and SlDEA1 CRISPR-edited plants were phenotypically better responsive to multiple stress factors taken under the study. The CRISPR-edited SlHyPRP1 and SlDEA1 plants showed a higher level of chlorophyll and proline content compared to wild-type (WT) plants under abiotic stress conditions. Reactive oxygen species accumulation and the cell death count per total area of leaves and roots under biotic stress were less in CRISPR-edited SlHyPRP1 and SlDEA1 plants compared to WT plants. The study reveals that the combined loss-of-function of SlHyPRP1 along with SlDEA1 is essential for imparting significant multi-stress tolerance (drought, salinity, bacterial leaf spot, and bacterial wilt) in S. lycopersicum L. The main feature of the study is the detailed genetic characterization of SlDEA1, a poorly studied 8CM family gene in multi-stress tolerance, through the CRISPR/Cas9 gene editing system. The study revealed the key negative regulatory role of SlDEA1 that function together as an anchor gene with SlHyPRP1 in imparting multi-stress tolerance in S. lycopersicum L. It was interesting that the present study also showed that transient CRISPR/Cas9 editing events of SlHyPRP1 and SlDEA1 genes were successfully replicated in stably generated parent-genome-edited line (GEd0) and genome-edited first-generation lines (GEd1) of S. lycopersicum L. With these upshots, the study’s key findings demonstrate outstanding value in developing sustainable multi-stress tolerance in S. lycopersicum L. and other crops to cope with climate change.

Introduction: Mulberry bacterial wilt is a serious destructive soil-borne disease caused by a complex and diverse group of pathogenic bacteria. Given that the bacterial wilt has been reported to cause a serious damage to the yield and quality of mulberry, therefore, elucidation of its main pathogenic groups is essential in improving our understanding of this disease and for the development of its potential control measures. Methods: In this study, combined metagenomic sequencing and culturedependent approaches were used to investigate the microbiome of healthy and bacterial wilt mulberry samples. Results: The results showed that the healthy samples had higher bacterial diversity compared to the diseased samples. Meanwhile, the proportion of opportunistic pathogenic and drug-resistant bacterial flora represented by Acinetobacter in the diseased samples was increased, while the proportion of beneficial bacterial flora represented by Proteobacteria was decreased. Ralstonia solanacearum species complex (RSSC), Enterobacter cloacae complex (ECC), Klebsiella pneumoniae, K. quasipneumoniae, K. michiganensis, K. oxytoca, and P. ananatis emerged as the main pathogens of the mulberry bacterial wilt. Discussion: In conclusion, this study provides a valuable reference for further focused research on the bacterial wilt of mulberry and other plants. [ABSTRACT FROM AUTHOR]

Bacterial wilt caused by Ralstonia solanacearum ranks the second top important bacterial plant disease worldwide. It is also the most important bacterial disease threatening the healthy development of Casuarina equisetifolia protection forest. 3-hydroxypalmitic acid methyl ester (3-OH PAME) functions as an important quorum sensing (QS) signal regulating the expression of virulence genes in R. solanacearum, and has been regarded as an ideal target for disease prevention and control. To screen native microorganisms capable of degrading 3-OH PAME, samples of C. equisetifolia branches and forest soil were collected and cultured in the medium containing 3-OH PAME as the sole carbon source. Bacteria with over 85% degradation rates of 3-OH PAME after 7-day incubation were further separated and purified. As a result, strain Q1-7 isolated from forest soil and strain Q4-3 isolated from C. equisetifolia branches were obtained and identified as Pseudomonas novel species Pseudomonas forestsoilum sp. nov. and P. tohonis, respectively, according to whole genome sequencing results. The degradation efficiencies of 3-OH PAME of strains Q1-7 and Q4-3 were 95.80% and 100.00% at 48 h, respectively. Both strains showed high esterase activities and inhibited R. solanacearum exopolysaccharide (EPS) and cellulase production. Application of strains Q1-7 and Q4-3 effectively protects C. equisetifolia, peanut and tomato plants from infection by R. solanacearum. Findings in this study provide potential resources for the prevention and control of bacterial wilt caused by R. solanacearum, as well as valuable materials for the identification of downstream quenching genes and the research and development of quenching enzymes for disease control. [ABSTRACT FROM AUTHOR]

Background: Tobacco bacterial wilt (TBW) and black shank (TBS) are responsible for substantial economic losses worldwide; however, microbial interactions and metabolisms in response to TBW and TBS pathogens in the tobacco rhizosphere remain unclear. Methods: We explored and compared the response of rhizosphere microbial communities to these two plant diseases with the incidences in moderate and heavy degrees by sequencing of 16S rRNA gene amplicons and bioinformatics analysis. Results and discussions: We found that the structure of rhizosphere soil bacterial communities was significantly (p < 0.05) changed from the incidences of TBW and TBS, which also led to decreased Shannon diversity and Pielou evenness. Compared with the healthy group (CK), the OTUs with significantly (p < 0.05) decreased relative abundances were mostly affiliated with Actinobacteria (e.g., Streptomyces and Arthrobacter) in the diseased groups, and the OTUs with significantly (p < 0.05) increased relative abundances were mainly identified as Proteobacteria and Acidobacteria. Also, molecular ecological network analysis showed that the nodes (<467) and links (<641) were decreased in the diseased groups compared with the control group (572; 1056), suggesting that both TBW and TBS weakened bacterial interactions. In addition, the predictive functional analysis indicated that the relative abundance of genes related to the biosynthesis of antibiotics (e.g., ansamycins and streptomycin) was significantly (p < 0.05) decreased due to incidences of TBW and TBS, and antimicrobial tests showed that some Actinobacteria strains (e.g., Streptomyces) and their secreted antibiotics (e.g., streptomycin) could effectively inhibit the growth of these two pathogens. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum is the causative agent of bacterial wilt of potato and other vegetable crops. Contaminated irrigation water contributes to the dissemination of this pathogen but the exact concentration or biological threshold to cause an infection is unknown. In two greenhouse experiments, potted potato plants (Solanum tuberosum) were exposed to a single irrigation with 50 mL water (non-invasive soil-soak inoculation) containing no or 10² – 108 CFU/mL R. solanacearum. The disease response of two cultivars, Kondor and HB, were compared. Disease development was monitored over a three-month period after which stems, roots and tubers of asymptomatic plants were analyzed for latent infections. First wilting symptoms were observed 15 days post inoculation in a plant inoculated with 5x109 CFU and a mean disease index was used to monitor disease development over time. An inoculum of 5x105 CFU per pot (1.3x10² CFU/g soil) was the minimum dose required to cause wilting symptoms, while one latent infection was detected at the lowest dose of 5x10² CFU per pot (0.13 CFU/g). In a second set of experiments, stem-inoculated potato plants grown in vitro were used to investigate the dose-response relationship under optimal conditions for pathogen growth and disease development. Plants were inoculated with doses between 0.5 and 5x105 CFU/plant which resulted in visible symptoms at all doses. The results led to a dose-response model describing the relationship between R. solanacearum exposure and probability of infection or illness of potato plants. Cultivar Kondor was more susceptible to brown-rot infections than HB in greenhouse experiments while there was no significant difference between the dose-response models of both cultivars in in vitro experiments. The ED50 for infection of cv Kondor was 1.1x107 CFU. Results can be used in management strategies aimed to reduce or eliminate the risk of bacterial wilt infection when using treated water in irrigation. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum is the causative agent of bacterial wilt of potato and other vegetable crops. Contaminated irrigation water contributes to the dissemination of this pathogen but the exact concentration or biological threshold to cause an infection is unknown. In two greenhouse experiments, potted potato plants (Solanum tuberosum) were exposed to a single irrigation with 50 mL water (non-invasive soil-soak inoculation) containing no or 102 – 108 CFU/mL R. solanacearum. The disease response of two cultivars, Kondor and HB, were compared. Disease development was monitored over a three-month period after which stems, roots and tubers of asymptomatic plants were analyzed for latent infections. First wilting symptoms were observed 15 days post inoculation in a plant inoculated with 5x109 CFU and a mean disease index was used to monitor disease development over time. An inoculum of 5x105 CFU per pot (1.3x102 CFU/g soil) was the minimum dose required to cause wilting symptoms, while one latent infection was detected at the lowest dose of 5x102 CFU per pot (0.13 CFU/g). In a second set of experiments, stem-inoculated potato plants grown in vitro were used to investigate the dose-response relationship under optimal conditions for pathogen growth and disease development. Plants were inoculated with doses between 0.5 and 5x105 CFU/plant which resulted in visible symptoms at all doses. The results led to a dose-response model describing the relationship between R. solanacearum exposure and probability of infection or illness of potato plants. Cultivar Kondor was more susceptible to brown-rot infections than HB in greenhouse experiments while there was no significant difference between the dose-response models of both cultivars in in vitro experiments. The ED50 for infection of cv Kondor was 1.1x107 CFU. Results can be used in management strategies aimed to reduce or eliminate the risk of bacterial wilt infection when using treated water in irrigation.

As one of the most important and largest transcription factors, WRKY plays a critical role in plant disease resistance. However, little is known regarding the functions of the WRKY family in cultivated peanuts (Arachis hypogaea L.). In this study, a total of 174 WRKY genes (AhWRKY) were identified from the genome of cultivated peanuts. Phylogenetic analysis revealed that AhWRKY proteins could be divided into four groups, including 35 (20.12%) in group I, 107 (61.49%) in group II, 31 (17.82%) in group III, and 1 (0.57%) in group IV. This division is further supported by the conserved motif compositions and intron/exon structures. All AhWRKY genes were unevenly located on all 20 chromosomes, among which 132 pairs of fragment duplication and seven pairs of tandem duplications existed. Eighteen miRNAs were found to be targeting 50 AhWRKY genes. Most AhWRKY genes fromsome groups showed tissue-specific expression. AhWRKY46, AhWRKY94, AhWRKY156, AhWRKY68, AhWRKY41, AhWRKY128, AhWRKY104, AhWRKY19, AhWRKY62, AhWRKY155, AhWRKY170, AhWRKY78, AhWRKY34, AhWRKY12, AhWRKY95, and AhWRKY76 were upregulated in ganhua18 and kainong313 genotypes after Ralstonia solanacearum infection. Ten AhWRKY genes (AhWRKY34, AhWRKY76, AhWRKY78, AhWRKY120, AhWRKY153, AhWRKY155, AhWRKY159, AhWRKY160, AhWRKY161, and AhWRKY162) from group III displayed di􀀀erent expression patterns in R. solanacearum sensitive and resistant peanut genotypes infected with the R. solanacearum. Two AhWRKY genes (AhWRKY76 and AhWRKY77) from group III obtained the LRR domain. AhWRKY77 downregulated in both genotypes; AhWRKY76 showed lower-higher expression in ganhua18 and higher expression in kainong313. Both AhWRKY76 and AhWRKY77 are targeted by ahy-miR3512, which may have an important function in peanut disease resistance. This study identified candidate WRKY genes with possible roles in peanut resistance against R. solanacearum infection. These findings not only contribute to our understanding of the novel role of WRKY family genes but also provide valuable information for disease resistance in A. hypogaea. [ABSTRACT FROM AUTHOR]

Bacterial wilt (BW) caused by Ralstonia solanacearum (R. solanacearum), is a vascular disease affecting diverse solanaceous crops and causing tremendous damage to crop production. However, our knowledge of the mechanism underlying its resistance or susceptibility is very limited. In this study, we characterized the physiological differences and compared the defense-related transcriptomes of two tobacco varieties, 4411-3 (highly resistant, HR) and K326 (moderately resistant, MR), after R. solanacearum infection at 0, 10, and 17 days after inoculation (dpi). A total of 3967 differentially expressed genes (DEGs) were identified between the HR and MR genotypes under mock condition at three time points, including1395 up-regulated genes in the HR genotype and 2640 up-regulated genes in the MR genotype. Also, 6,233 and 21,541 DEGs were induced in the HR and MR genotypes after R. solanacearum infection, respectively. Furthermore, GO and KEGG analyses revealed that DEGs in the HR genotype were related to the cell wall, starch and sucrose metabolism, glutathione metabolism, ABC transporters, endocytosis, glycerolipid metabolism, and glycerophospholipid metabolism. The defense-related genes generally showed genotype-specific regulation and expression differences after R. solanacearum infection. In addition, genes related to auxin and ABA were dramatically up-regulated in the HR genotype. The contents of auxin and ABA in the MR genotype were significantly higher than those in the HR genotype after R. solanacearum infection, providing insight into the defense mechanisms of tobacco. Altogether, these results clarify the physiological and transcriptional regulation of R. solanacearum resistance infection in tobacco, and improve our understanding of the molecular mechanism underlying the plant-pathogen interaction. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum causes bacterial wilt, a devastating plant disease, responsible for serious losses on many crop plants. R. solanacearum phylotype II-B1 strains have caused important outbreaks in temperate regions, where the pathogen has been identified inside asymptomatic bittersweet (Solanum dulcamara) plants near rivers and in potato fields. S. dulcamara is a perennial species described as a reservoir host where R. solanacearum can overwinter, but their interaction remains uncharacterised. In this study, we have systematically analysed R. solanacearum infection in S. dulcamara , dissecting the behaviour of this plant compared with susceptible hosts such as tomato cv. Marmande, for which the interaction is well described. Compared with susceptible tomatoes, S. dulcamara plants (i) show delayed symptomatology and bacterial progression, (ii) restrict bacterial movement inside and between xylem vessels, (iii) limit bacterial root colonisation, and (iv) show constitutively higher lignification in the stem. Taken together, these results demonstrate that S. dulcamara behaves as partially resistant to bacterial wilt, a property that is enhanced at lower temperatures. This study proves that tolerance (i.e., the capacity to reduce the negative effects of infection) is not required for a wild plant to act as a reservoir host. We propose that inherent resistance (impediment to colonisation) and a perennial habit enable bittersweet plants to behave as reservoirs for R. solanacearum. [ABSTRACT FROM AUTHOR]

Bacterial wilt (BW) caused by Ralstonia solanacearum (R. solanacearum), is a vascular disease affecting diverse solanaceous crops and causing tremendous damage to crop production. However, our knowledge of the mechanism underlying its resistance or susceptibility is very limited. In this study, we characterized the physiological differences and compared the defense-related transcriptomes of two tobacco varieties, 4411-3 (highly resistant, HR) and K326 (moderately resistant, MR), after R. solanacearum infection at 0, 10, and 17 days after inoculation (dpi). A total of 3967 differentially expressed genes (DEGs) were identified between the HR and MR genotypes under mock condition at three time points, including1395 up-regulated genes in the HR genotype and 2640 up-regulated genes in the MR genotype. Also, 6,233 and 21,541 DEGs were induced in the HR and MR genotypes after R. solanacearum infection, respectively. Furthermore, GO and KEGG analyses revealed that DEGs in the HR genotype were related to the cell wall, starch and sucrose metabolism, glutathione metabolism, ABC transporters, endocytosis, glycerolipid metabolism, and glycerophospholipid metabolism. The defense-related genes generally showed genotype-specific regulation and expression differences after R. solanacearum infection. In addition, genes related to auxin and ABA were dramatically up-regulated in the HR genotype. The contents of auxin and ABA in the MR genotype were significantly higher than those in the HR genotype after R. solanacearum infection, providing insight into the defense mechanisms of tobacco. Altogether, these results clarify the physiological and transcriptional regulation of R. solanacearum resistance infection in tobacco, and improve our understanding of the molecular mechanism underlying the plant-pathogen interaction.

Ralstonia solanacearum causes bacterial wilt, a devastating plant disease, responsible for serious losses on many crop plants. R. solanacearum phylotype II-B1 strains have caused important outbreaks in temperate regions, where the pathogen has been identified inside asymptomatic bittersweet (Solanum dulcamara) plants near rivers and in potato fields. S. dulcamara is a perennial species described as a reservoir host where R. solanacearum can overwinter, but their interaction remains uncharacterised. In this study, we have systematically analysed R. solanacearum infection in S. dulcamara, dissecting the behaviour of this plant compared with susceptible hosts such as tomato cv. Marmande, for which the interaction is well described. Compared with susceptible tomatoes, S. dulcamara plants (i) show delayed symptomatology and bacterial progression, (ii) restrict bacterial movement inside and between xylem vessels, (iii) limit bacterial root colonisation, and (iv) show constitutively higher lignification in the stem. Taken together, these results demonstrate that S. dulcamara behaves as partially resistant to bacterial wilt, a property that is enhanced at lower temperatures. This study proves that tolerance (i.e., the capacity to reduce the negative effects of infection) is not required for a wild plant to act as a reservoir host. We propose that inherent resistance (impediment to colonisation) and a perennial habit enable bittersweet plants to behave as reservoirs for R. solanacearum.

Plant-associated microbiota plays an important role in plant disease resistance. Bacterial wilt resistance of tomato is a function of the quantitative trait of tomato plants; however, the mechanism underlying quantitative resistance is unexplored. In this study, we hypothesized that rhizosphere microbiota affects the resistance of tomato plants against soil-borne bacterial wilt caused by Ralstonia solanacearum. This hypothesis was tested using a tomato cultivar grown in a defined soil with various microbiota transplants. The bacterial wilt-resistant Hawaii 7996 tomato cultivar exhibited marked suppression and induction of disease severity after treatment with upland soil-derived and forest soil-derived microbiotas, respectively, whereas the transplants did not affect the disease severity in the susceptible tomato cultivar Moneymaker. The differential resistance of Hawaii 7996 to bacterial wilt was abolished by diluted or heat-killed microbiota transplantation. Microbial community analysis revealed the transplant-specific distinct community structure in the tomato rhizosphere and the significant enrichment of specific microbial operational taxonomic units (OTUs) in the rhizosphere of the upland soil microbiota-treated Hawaii 7996. These results suggest that the specific transplanted microbiota alters the bacterial wilt resistance in the resistant cultivar potentially through a priority effect.

Ralstonia solanacearum is the causative agent of bacterial wilt disease on a wide range of plant species. Besides the numerous bacterial activities required for host invasion, those involved in the adaptation to the plant environment are key for the success of infection. R. solanacearum ability to cope with the oxidative burst produced by the plant is likely one of the activities required to grow parasitically. Among the multiple reactive oxygen species (ROS)-scavenging enzymes predicted in the R. solanacearum GMI1000 genome, a single monofunctional catalase (KatE) and two KatG bifunctional catalases were identified. In this work, we show that these catalase activities are active in bacterial protein extracts and demonstrate by gene disruption and mutant complementation that the monofunctional catalase activity is encoded by katE. Different strategies were used to evaluate the role of KatE in bacterial physiology and during the infection process that causes bacterial wilt. We show that the activity of the enzyme is maximal during exponential growth in vitro and this growth-phase regulation occurs at the transcriptional level. Our studies also demonstrate that katE expression is transcriptionally activated by HrpG, a central regulator of R. solanacearum induced upon contact with the plant cells. In addition, we reveal that even though both KatE and KatG catalase activities are induced upon hydrogen peroxide treatment, KatE has a major effect on bacterial survival under oxidative stress conditions and especially in the adaptive response of R. solanacearum to this oxidant. The katE mutant strain also exhibited differences in the structural characteristics of the biofilms developed on an abiotic surface in comparison to wild-type cells, but not in the overall amount of biofilm production. The role of catalase KatE during the interaction with its host plant tomato is also studied, revealing that disruption of this gene has no effect on R. solanacearum virulence or bacterial growth in leave tissues, which suggests a minor role for this catalase in bacterial fitness in planta. Our work provides the first characterization of the R. solanacearum catalases and identifies KatE as a bona fide monofunctional catalase with an important role in bacterial protection against oxidative stress.

Plant-associated microbiota plays an important role in plant disease resistance. Bacterial wilt resistance of tomato is a function of the quantitative trait of tomato plants; however, the mechanism underlying quantitative resistance is unexplored. In this study, we hypothesized that rhizosphere microbiota affects the resistance of tomato plants against soil-borne bacterial wilt caused by Ralstonia solanacearum. This hypothesis was tested using a tomato cultivar grown in a defined soil with various microbiota transplants. The bacterial wilt-resistant Hawaii 7996 tomato cultivar exhibited marked suppression and induction of disease severity after treatment with upland soil-derived and forest soil-derived microbiotas, respectively, whereas the transplants did not affect the disease severity in the susceptible tomato cultivar Moneymaker. The differential resistance of Hawaii 7996 to bacterial wilt was abolished by diluted or heat-killed microbiota transplantation. Microbial community analysis revealed the transplant-specific distinct community structure in the tomato rhizosphere and the significant enrichment of specific microbial operational taxonomic units (OTUs) in the rhizosphere of the upland soil microbiota-treated Hawaii 7996. These results suggest that the specific transplanted microbiota alters the bacterial wilt resistance in the resistant cultivar potentially through a priority effect. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum is the causative agent of bacterial wilt disease on a wide range of plant species. Besides the numerous bacterial activities required for host invasion, those involved in the adaptation to the plant environment are key for the success of infection. R. solanacearum ability to cope with the oxidative burst produced by the plant is likely one of the activities required to grow parasitically. Among the multiple reactive oxygen species (ROS)-scavenging enzymes predicted in the R. solanacearum GMI1000 genome, a single monofunctional catalase (KatE) and two KatG bifunctional catalases were identified. In this work, we show that these catalase activities are active in bacterial protein extracts and demonstrate by gene disruption and mutant complementation that the monofunctional catalase activity is encoded by katE. Different strategies were used to evaluate the role of KatE in bacterial physiology and during the infection process that causes bacterial wilt. We show that the activity of the enzyme is maximal during exponential growth in vitro and this growth-phase regulation occurs at the transcriptional level. Our studies also demonstrate that katE expression is transcriptionally activated by HrpG, a central regulator of R. solanacearum induced upon contact with the plant cells. In addition, we reveal that even though both KatE and KatG catalase activities are induced upon hydrogen peroxide treatment, KatE has a major effect on bacterial survival under oxidative stress conditions and especially in the adaptive response of R. solanacearum to this oxidant. The katE mutant strain also exhibited differences in the structural characteristics of the biofilms developed on an abiotic surface in comparison to wild-type cells, but not in the overall amount of biofilm production. The role of catalase KatE during the interaction with its host plant tomato is also studied, revealing that disruption of this gene has no effect on R. solanacearum virulence or bacterial growth in leave tissues, which suggests a minor role for this catalase in bacterial fitness in planta. Our work provides the first characterization of the R. solanacearum catalases and identifies KatE as a bona fide monofunctional catalase with an important role in bacterial protection against oxidative stress. [ABSTRACT FROM AUTHOR]

Understanding the response of soil microbial communities to pathogenic Ralstonia solanacearum is crucial for preventing bacterial wilt outbreaks. In this study, we investigated the soil physicochemical and microbial community to assess their impact on the pathogenic R.solanacearum through metagenomics. Our results revealed that certain archaeal taxa were the main contributors influencing the health of plants. Additionally, the presence of the pathogen showed a strong negative correlation with soil phosphorus levels, while soil phosphorus was significantly correlated with bacterial and archaeal communities. We found that the network of microbial interactions in healthy plant rhizosphere soils was more complex compared to diseased soils. The diseased soil network had more linkages, particularly related to the pathogen occurrence. Within the network, the family Comamonadaceae, specifically Ramlibacter&#95;tataouinensis , was enriched in healthy samples and showed a significantly negative correlation with the pathogen. In terms of archaea, Halorubrum , Halorussus&#95;halophilus (family: Halobacteriaceae), and Natronomonas&#95;pharaonis (family: Haloarculaceae) were enriched in healthy plant rhizosphere soils and showed negative correlations with R.solanacearum . These findings suggested that the presence of these archaea may potentially reduce the occurrence of bacterial wilt disease. On the other hand, Halostagnicola&#95;larseniia and Haloterrigena &#95;sp.&#95;BND6 (family: Natrialbaceae) had higher relative abundance in diseased plants and exhibited significantly positive correlations with R.solanacearum , indicating their potential contribution to the pathogen's occurrence. Moreover, we explored the possibility of functional gene sharing among the correlating bacterial pairs within the Molecular Ecological Network. Our analysis revealed 468 entries of horizontal gene transfer (HGT) events, emphasizing the significance of HGT in shaping the adaptive traits of plant-associated bacteria, particularly in relation to host colonization and pathogenicity. Overall, this work revealed key factors, patterns and response mechanisms underlying the rhizosphere soil microbial populations. The findings offer valuable guidance for effectively controlling soil-borne bacterial diseases and developing sustainable agriculture practices., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Xiao, Zhang, Li, Teng, Wu, Liu, Yu, He, Li, Li, Meng, Yin and Wang.)

The soil microbial community plays a critical role in promoting robust plant growth and serves as an effective defence mechanism against root pathogens. Current research has focused on unravelling the compositions and functions of diverse microbial taxa in plant rhizospheres invaded by Ralstonia solanacearum , however, the specific mechanisms by which key microbial groups with distinct functions exert their effects remain unclear. In this study, we employed a combination of amplicon sequencing and metabolomics analysis to investigate the principal metabolic mechanisms of key microbial taxa in plant rhizosphere soil. Compared to the healthy tobacco rhizosphere samples, the bacterial diversity and co-occurrence network of the diseased tobacco rhizosphere soil were significantly reduced. Notably, certain genera, including Gaiella , Rhodoplanes , and MND1 ( Nitrosomonadaceae ), were found to be significantly more abundant in the rhizosphere of healthy plants than in that of diseased plants. Eight environmental factors, including exchangeable magnesium, available phosphorus, and pH, were found to be crucial factors influencing the composition of the microbial community. Ralstonia displayed negative correlations with pH, exchangeable magnesium, and cation exchange flux, but showed a positive correlation with available iron. Furthermore, metabolomic analysis revealed that the metabolic pathways related to the synthesis of various antibacterial compounds were significantly enriched in the healthy group. The correlation analysis results indicate that the bacterial genera Polycyclovorans , Lysobacter , Pseudomonas , and Nitrosospira may participate in the synthesis of antibacterial compounds. Collectively, our findings contribute to a more in-depth understanding of disease resistance mechanisms within healthy microbial communities and provide a theoretical foundation for the development of targeted strategies using beneficial microorganisms to suppress disease occurrence., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Wei, Liang, Wang, Chi, Wang, Tan, Shi, Song, Cui, Xie, Cheng and Wang.)

Ralstonia solanacearum species complex is a devastating phytopathogen with an unusually wide host range, and new host plants are continuously being discovered. In June 2016, a new bacterial wilt on Cucurbita maxima was observed in Guangdong province, China. Initially, in the adult plant stage, several leaves of each plant withered suddenly and drooped; the plant then wilted completely, and the color of their vasculature changed to dark brown, ultimately causing the entire plant to die. Creamy-whitish bacterial masses were observed to ooze from crosscut stems of these diseased plants. To develop control strategies for C. maxima bacterial wilt, the causative pathogenic isolates were identified and characterized. Twenty-four bacterial isolates were obtained from diseased C. maxima plants, and 16S rRNA gene sequencing and pathogenicity analysis results indicated that the pathogen of C. maxima bacterial wilt was Ralstonia solanacearum. The results from DNA-based analysis, host range determination and bacteriological identification confirmed that the 24 isolates belonged to R. solanacearum phylotype I, race 1, and eight of these isolates belonged to biovar 3, while 16 belonged to biovar 4. Based on the results of partial egl gene sequence analysis, the 24 isolates clustered into three egl- sequence type groups, sequevars 17, 45, and 56. Sequevar 56 is a new sequevar which is described for the first time in this paper. An assessment of the resistance of 21 pumpkin cultivars revealed that C. moschata cv. Xiangyu1 is resistant to strain RS378, C. moschata cv. Xiangmi is moderately resistant to strain RS378, and 19 other pumpkin cultivars, including four C. maxima cultivars and 15 C. moschata cultivars, are susceptible to strain RS378. To the best of our knowledge, this is the first report of C. maxima bacterial wilt caused by R. solanacearum race 1 in the world. Our results provide valuable information for the further development of control strategies for C. maxima wilt disease.

Bacterial wilt (BW) caused by Ralstonia solanacearum is responsible for substantial losses in cultivated potato (Solanum tuberosum) crops worldwide. Resistance genes have been identified in wild species; however, introduction of these through classical breeding has achieved only partial resistance, which has been linked to poor agronomic performance. The Arabidopsis thaliana (At) pattern recognition receptor elongation factor-Tu (EF-Tu) receptor (EFR) recognizes the bacterial pathogen-associated molecular pattern EF-Tu (and its derived peptide elf18) to confer anti-bacterial immunity. Previous work has shown that transfer of AtEFR into tomato confers increased resistance to R. solanacearum. Here, we evaluated whether the transgenic expression of AtEFR would similarly increase BW resistance in a commercial potato line (INIA Iporá), as well as in a breeding potato line (09509.6) in which quantitative resistance has been introgressed from the wild potato relative Solanum commersonii. Resistance to R. solanacearum was evaluated by damaged root inoculation under controlled conditions. Both INIA Iporá and 09509.6 potato lines expressing AtEFR showed greater resistance to R. solanacearum, with no detectable bacteria in tubers evaluated by multiplex-PCR and plate counting. Notably, AtEFR expression and the introgression of quantitative resistance from S. commersonii had a significant additive effect in 09509.6-AtEFR lines. These results show that the combination of heterologous expression of AtEFR with quantitative resistance introgressed from wild relatives is a promising strategy to develop BW resistance in potato.

Bacterial wilt caused by plant pathogenic Ralstonia spp. is one of the most important diseases affecting the production of many important crops worldwide. In China, a large scientific community has been dedicated to studying bacterial wilt and its causative agent, Ralstonia pseudosolanacearum and R. solanacearum. Most of their work was published in Chinese, which has hindered international communication and collaboration in this field. In this review, we summarize the status of knowledge on geographical distribution, diversity, and host range of Ralstonia spp., as well as, the impact of bacterial wilt on important crops and disease control approaches, in China. We present areas of research and publications by Chinese scientists and propose the promotion of collaborative research within China and with the international community.

Potato (Solanum tuberosum L.) is one of the main hosts of Ralstonia solanacearum, the causative agent of bacterial wilt. This plant pathogen bacteria produce asymptomatic latent infections that promote its global spread, hindering disease control. A potato breeding program is conducted in Uruguay based on the introgression of resistance from the wild native species S. commersonii Dun. Currently, several backcrosses were generated exploiting the high genetic variability of this wild species resulting in advanced interspecific breeding lines with different levels of bacterial wilt resistance. The overall aim of this work was to characterize the interaction of the improved potato germplasm with R. solanacearum. Potato clones with different responses to R. solanacearum were selected, and colonization, dissemination and multiplication patterns after infection were evaluated. A R. solanacearum strain belonging to the phylotype IIB-sequevar 1, with high aggressiveness on potato was genetically modified to constitutively generate fluorescence and luminescence from either the green fluorescence protein gene or lux operon. These reporter strains were used to allow a direct and precise visualization of fluorescent and luminescent cells in plant tissues by confocal microscopy and luminometry. Based on wilting scoring and detection of latent infections, the selected clones were classified as susceptible or tolerant, while no immune-like resistance response was identified. Typical wilting symptoms in susceptible plants were correlated with high concentrations of bacteria in roots and along the stems. Tolerant clones showed a colonization pattern restricted to roots and a limited number of xylem vessels only in the stem base. Results indicate that resistance in potato is achieved through restriction of bacterial invasion and multiplication inside plant tissues, particularly in stems. Tolerant plants were also characterized by induction of anatomical and biochemical changes after R. solanacearum infection, including hyperplasic activity of conductor tissue, tylose production, callose and lignin deposition, and accumulation of reactive oxygen species. This study highlights the potential of the identified tolerant interspecific potato clones as valuable genetic resources for potato-breeding programs and leads to a better understanding of resistance against R. solanacearum in potato.

The current bacterial wilt infestation level in the potato fields in the Peruvian Andes was investigated by collecting stem samples from wilted plants and detecting Ralstonia solanacearum. In total 39 farmers’ fields located in the central and northern Peru between the altitudes 2111 and 3742 m above sea level were sampled. R. solanacearum was detected in 19 fields, and in 153 out of the 358 samples analyzed. Phylogenetic analysis using the partial sequence of the endoglucanase gene on strains collected in Peru between 1966 and 2016 from potato, pepper, tomato, plantain or soil, divided the strains in phylotypes I, IIA, and IIB. The Phylotype IIB isolates formed seven sequevar groups including the previously identified sequevars 1, 2, 3, 4, and 25. In addition to this, three new sequevars of phylotype IIB were identified. Phylotype IIA isolates from Peru clustered together with reference strains previously assigned to sequevars 5, 39, 41, and 50, and additionally one new sequevar was identified. The Phylotype I strain was similar to the sequevar 18. Most of the Peruvian R. solanacearum isolates were IIB-1 strains. In the old collection sampled between 1966 and 2013, 72% were IIB-1 and in the new collection at 2016 no other strains were found. The pathogenicity of 25 isolates representing the IIA and IIB sequevar groups was tested on potato, tomato, eggplant and tobacco. All were highly aggressive on potato, but differed in pathogenicity on the other hosts, especially on tobacco. All IIA strains caused latent infection on tobacco and some strains also caused wilting, while IIB strains caused only few latent infections on this species. In conclusion, high molecular diversity was found among the R. solanacearum strains in Peru. Most of the variability was found in areas that are no longer used for potato cultivation and thus these strains do not pose a real threat for potato production in the country. Compared to the previous data from the 1990s, the incidence of bacterial wilt has decreased in Peru. The epidemics are likely caused by infected seed tubers carrying the clonal brown rot strain IIB-1.

Ralstonia solanacearum and R. pseudosolanacearum are well known primary pathogens of herbaceous crops. Reports of wilt caused by these pathogens in tree species are limited other than on Eucalyptus species. Despite the widespread occurrence of so-called bacterial wilt on eucalypts in tropical and sub-tropical parts of Africa, Asia, and the Americas, there remain many contradictions relating to the disease. Our field observations over many years in most regions where the disease occurs on Eucalyptus show that it is always associated with trees that have been subjected to severe stress. The disease is typically diagnosed by immersing cut stems in water and observing bacterial streaming, but the identity of the bacteria within this suspension is seldom considered. To add to the confusion, pathogenicity tests on susceptible species or clones are rarely successful. When they do work, they are on small plants in greenhouse trials. It has become all to easy to attribute Eucalyptus death exclusively to Ralstonia infection. Our data strongly suggest that Ralstonia species and probably other bacteria are latent colonists commonly occurring in healthy and particularly clonally propagated eucalypts. The onset of stress factors provide the bacteria with an opportunity to develop. We believe that the resulting stress weakens the defense systems of the trees allowing Ralstonia and bacterial endophytes to proliferate. Overall our research suggests that R. solanacearum and R. pseudosolanacearum are not primary pathogens of Eucalyptus. Short of clear evidence that they are primary pathogens of Eucalyptus it is inappropriate to attribute this disease solely to infection by Ralstonia species.

Bacterial wilt (BW) is a major disease of solanaceous crops caused by the Ralstonia solanacearum species complex (RSSC). Strains are grouped into five phylotypes (I, IIA, IIB, III, and IV). Varietal resistance is the most sustainable strategy for managing BW. Nevertheless, breeding to improve cultivar resistance has been limited by the pathogen’s extensive genetic diversity. Identifying the genetic bases of specific and non-specific resistance is a prerequisite to breed improvement. A major gene (ERs1) was previously mapped in eggplant (Solanum melongena L.) using an intraspecific population of recombinant inbred lines derived from the cross of susceptible MM738 (S) × resistant AG91-25 (R). ERs1 was originally found to control three strains from phylotype I, while being totally ineffective against a virulent strain from the same phylotype. We tested this population against four additional RSSC strains, representing phylotypes I, IIA, IIB, and III in order to clarify the action spectrum of ERs1. We recorded wilting symptoms and bacterial stem colonization under controlled artificial inoculation. We constructed a high-density genetic map of the population using single nucleotide polymorphisms (SNPs) developed from genotyping-by-sequencing and added 168 molecular markers [amplified fragment length polymorphisms (AFLPs), simple sequence repeats (SSRs), and sequence-related amplified polymorphisms (SRAPs)] developed previously. The new linkage map based on a total of 1,035 markers was anchored on eggplant, tomato, and potato genomes. Quantitative trait locus (QTL) mapping for resistance against a total of eight RSSC strains resulted in the detection of one major phylotype-specific QTL and two broad-spectrum QTLs. The major QTL, which specifically controls three phylotype I strains, was located at the bottom of chromosome 9 and corresponded to the previously identified major gene ERs1. Five candidate R-genes were underlying this QTL, with different alleles between the parents. The two other QTLs detected on chromosomes 2 and 5 were found to be associated with partial resistance to strains of phylotypes I, IIA, III and strains of phylotypes IIA and III, respectively. Markers closely linked to these three QTLs will be crucial for breeding eggplant with broad-spectrum resistance to BW. Furthermore, our study provides an important contribution to the molecular characterization of ERs1, which was initially considered to be a major resistance gene.

Bacterial wilt (BW) caused by Ralstonia solanacearum is responsible for substantial losses in cultivated potato (Solanum tuberosum) crops worldwide. Resistance genes have been identified in wild species; however, introduction of these through classical breeding has achieved only partial resistance, which has been linked to poor agronomic performance. The Arabidopsis thaliana (At) pattern recognition receptor elongation factor-Tu (EF-Tu) receptor (EFR) recognizes the bacterial pathogen-associated molecular pattern EF-Tu (and its derived peptide elf18) to confer anti-bacterial immunity. Previous work has shown that transfer of AtEFR into tomato confers increased resistance to R. solanacearum. Here, we evaluated whether the transgenic expression of AtEFR would similarly increase BW resistance in a commercial potato line (INIA Iporá), as well as in a breeding potato line (09509.6) in which quantitative resistance has been introgressed from the wild potato relative Solanum commersonii. Resistance to R. solanacearum was evaluated by damaged root inoculation under controlled conditions. Both INIA Iporá and 09509.6 potato lines expressing AtEFR showed greater resistance to R. solanacearum, with no detectable bacteria in tubers evaluated by multiplex-PCR and plate counting. Notably, AtEFR expression and the introgression of quantitative resistance from S. commersonii had a significant additive effect in 09509.6-AtEFR lines. These results show that the combination of heterologous expression of AtEFR with quantitative resistance introgressed from wild relatives is a promising strategy to develop BW resistance in potato. [ABSTRACT FROM AUTHOR]

Potato (Solanum tuberosum L.) is one of the main hosts of Ralstonia solanacearum, the causative agent of bacterial wilt. This plant pathogen bacteria produce asymptomatic latent infections that promote its global spread, hindering disease control. A potato breeding program is conducted in Uruguay based on the introgression of resistance from the wild native species S. commersonii Dun. Currently, several backcrosses were generated exploiting the high genetic variability of this wild species resulting in advanced interspecific breeding lines with different levels of bacterial wilt resistance. The overall aim of this work was to characterize the interaction of the improved potato germplasm with R. solanacearum. Potato clones with different responses to R. solanacearum were selected, and colonization, dissemination and multiplication patterns after infection were evaluated. A R. solanacearum strain belonging to the phylotype IIB-sequevar 1, with high aggressiveness on potato was genetically modified to constitutively generate fluorescence and luminescence from either the green fluorescence protein gene or lux operon. These reporter strains were used to allow a direct and precise visualization of fluorescent and luminescent cells in plant tissues by confocal microscopy and luminometry. Based on wilting scoring and detection of latent infections, the selected clones were classified as susceptible or tolerant, while no immune-like resistance response was identified. Typical wilting symptoms in susceptible plants were correlated with high concentrations of bacteria in roots and along the stems. Tolerant clones showed a colonization pattern restricted to roots and a limited number of xylem vessels only in the stem base. Results indicate that resistance in potato is achieved through restriction of bacterial invasion and multiplication inside plant tissues, particularly in stems. Tolerant plants were also characterized by induction of anatomical and biochemical changes after R. solanacearum infection, including hyperplasic activity of conductor tissue, tylose production, callose and lignin deposition and accumulation of reactive oxygen species. This study highlights the potential of the identified tolerant interspecific potato clones as valuable genetic resources for potato-breeding programs and leads to a better understanding of resistance against R. solanacearum in potato. [ABSTRACT FROM AUTHOR]

The current bacterial wilt infestation level in the potato fields in the Peruvian Andes was investigated by collecting stem samples from wilted plants and detecting Ralstonia solanacearum. In total 39 farmers' fields located in the central and northern Peru between the altitudes 2111 and 3742 m above sea level were sampled. R. solanacearum was detected in 19 fields, and in 153 out of the 358 samples analyzed. Phylogenetic analysis using the partial sequence of the endoglucanase gene on strains collected in Peru between 1966 and 2016 from potato, pepper, tomato, plantain or soil, divided the strains in phylotypes I, IIA, and IIB. The Phylotype IIB isolates formed seven sequevar groups including the previously identified sequevars 1, 2, 3, 4, and 25. In addition to this, three new sequevars of phylotype IIB were identified. Phylotype IIA isolates from Peru clustered together with reference strains previously assigned to sequevars 5, 39, 41, and 50, and additionally one new sequevar was identified. The Phylotype I strain was similar to the sequevar 18. Most of the Peruvian R. solanacearum isolates were IIB-1 strains. In the old collection sampled between 1966 and 2013, 72% were IIB-1 and in the new collection at 2016 no other strains were found. The pathogenicity of 25 isolates representing the IIA and IIB sequevar groups was tested on potato, tomato, eggplant and tobacco. All were highly aggressive on potato, but differed in pathogenicity on the other hosts, especially on tobacco. All IIA strains caused latent infection on tobacco and some strains also caused wilting, while IIB strains caused only few latent infections on this species. In conclusion, high molecular diversity was found among the R. solanacearum strains in Peru. Most of the variability was found in areas that are no longer used for potato cultivation and thus these strains do not pose a real threat for potato production in the country. Compared to the previous data from the 1990s, the incidence of bacterial wilt has decreased in Peru. The epidemics are likely caused by infected seed tubers carrying the clonal brown rot strain IIB-1. [ABSTRACT FROM AUTHOR]

Bacterial wilt (BW) is a major disease of solanaceous crops caused by the Ralstonia solanacearum species complex (RSSC). Strains are grouped into five phylotypes (I, IIA, IIB, III, and IV). Varietal resistance is the most sustainable strategy for managing BW. Nevertheless, breeding to improve cultivar resistance has been limited by the pathogen's extensive genetic diversity. Identifying the genetic bases of specific and non-specific resistance is a prerequisite to breed improvement. A major gene (ERs1) was previously mapped in eggplant (Solanum melongena L.) using an intraspecific population of recombinant inbred lines derived from the cross of susceptible MM738 (S) x resistant AG91-25 (R). ERs1 was originally found to control three strains from phylotype I, while being totally ineffective against a virulent strain from the same phylotype. We tested this population against four additional RSSC strains, representing phylotypes I, IIA, IIB, and III in order to clarify the action spectrum of ERs1. We recorded wilting symptoms and bacterial stem colonization under controlled artificial inoculation. We constructed a high-density genetic map of the population using single nucleotide polymorphisms (SNPs) developed from genotyping-by-sequencing and added 168 molecular markers [amplified fragment length polymorphisms (AFLPs), simple sequence repeats (SSRs), and sequence-related amplified polymorphisms (SRAPs)] developed previously. The new linkage map based on a total of 1,035 markers was anchored on eggplant, tomato, and potato genomes. Quantitative trait locus (QTL) mapping for resistance against a total of eight RSSC strains resulted in the detection of one major phylotype-specific QTL and two broad-spectrum QTLs. The major QTL, which specifically controls three phylotype I strains, was located at the bottom of chromosome 9 and corresponded to the previously identified major gene ERs1. Five candidate R-genes were underlying this QTL, with different alleles between the parents. The two other QTLs detected on chromosomes 2 and 5 were found to be associated with partial resistance to strains of phylotypes I, IIA, III and strains of phylotypes IIA and III, respectively. Markers closely linked to these three QTLs will be crucial for breeding eggplant with broad-spectrum resistance to BW. Furthermore, our study provides an important contribution to the molecular characterization of ERs1, which was initially considered to be a major resistance gene. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum and R. pseudosolanacearum are well known primary pathogens of herbaceous crops. Reports of wilt caused by these pathogens in tree species are limited other than on Eucalyptus species. Despite the widespread occurrence of so-called bacterial wilt on eucalypts in tropical and sub-tropical parts of Africa, Asia, and the Americas, there remain many contradictions relating to the disease. Our field observations over many years in most regions where the disease occurs on Eucalyptus show that it is always associated with trees that have been subjected to severe stress. The disease is typically diagnosed by immersing cut stems in water and observing bacterial streaming, but the identity of the bacteria within this suspension is seldom considered. To add to the confusion, pathogenicity tests on susceptible species or clones are rarely successful. When they do work, they are on small plants in greenhouse trials. It has become all to easy to attribute Eucalyptus death exclusively to Ralstonia infection. Our data strongly suggest that Ralstonia species and probably other bacteria are latent colonists commonly occurring in healthy and particularly clonally propagated eucalypts. The onset of stress factors provide the bacteria with an opportunity to develop. We believe that the resulting stress weakens the defense systems of the trees allowing Ralstonia and bacterial endophytes to proliferate. Overall our research suggests that R. solanacearum and R. pseudosolanacearum are not primary pathogens of Eucalyptus. Short of clear evidence that they are primary pathogens of Eucalyptus it is inappropriate to attribute this disease solely to infection by Ralstonia species. [ABSTRACT FROM AUTHOR]

Bacterial wilt of potatoes--also called brown rot--is a devastating disease caused by the vascular pathogen Ralstonia solanacearum that leads to significant yield loss. As in other plant-pathogen interactions, the first contacts established between the bacterium and the plant largely condition the disease outcome. Here, we studied the transcriptome of R. solanacearum UY031 early after infection in two accessions of the wild potato Solanum commersonii showing contrasting resistance to bacterial wilt. Total RNAs obtained from asymptomatic infected roots were deep sequenced and for 4,609 out of the 4,778 annotated genes in strain UY031 were recovered. Only 2 genes were differentially-expressed between the resistant and the susceptible plant accessions, suggesting that the bacterial component plays a minor role in the establishment of disease. On the contrary, 422 genes were differentially expressed (DE) in planta compared to growth on a synthetic rich medium. Only 73 of these genes had been previously identified as DE in a transcriptome of R. solanacearum extracted from infected tomato xylem vessels. Virulence determinants such as the Type Three Secretion System (T3SS) and its effector proteins, motility structures, and reactive oxygen species (ROS) detoxifying enzymes were induced during infection of S. commersonii. On the contrary, metabolic activities were mostly repressed during early root colonization, with the notable exception of nitrogen metabolism, sulfate reduction and phosphate uptake. Several of the R. solanacearum genes identified as significantly up-regulated during infection had not been previously described as virulence factors. This is the first report describing the R. solanacearum transcriptome directly obtained from infected tissue and also the first to analyze bacterial gene expression in the roots, where plant infection takes place. We also demonstrate that the bacterial transcriptome in planta can be studied when pathogen numbers are low by sequencing transcripts from infected tissue avoiding prokaryotic RNA enrichment. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum phylotype I mulberry strains are causative agent of bacterial wilt of mulberry. Current diagnostic methods are not adopted to the mulberry wilt disease. In this study, we developed a rapid method, loop-mediated isothermal amplification (LAMP), to detect R. solanacearum phylotype I mulberry strains. A set of six primers was designed to target the clone MG67 sequence in this LAMP detection which can be completed in 20 min at 64°C. The results of the LAMP reaction could be observed with the naked eye due to magnesium pyrophosphate precipitate produced during the reaction or the color change after adding SYBR Green I. The specificity of the LAMP was confirmed using DNA from 46 representative strains of R. solanacearum and 7 other soil-borne bacteria strains. This method was also of high sensitivity and could be used to detect the presence of less than 160 fg genomic DNA or 2.2 x 10² CFU/ml of bacterial cells per 25 μl reaction volume, moreover, the presence of plant tissue fluid did not affect the sensitivity. Since it does not require expensive equipment or specialized techniques, this LAMP-based diagnostic method has the potential to be used under field conditions to make disease forecasting more accurate and efficient. [ABSTRACT FROM AUTHOR]

Introduction: Mulberry bacterial wilt is a serious destructive soil-borne disease caused by a complex and diverse group of pathogenic bacteria. Given that the bacterial wilt has been reported to cause a serious damage to the yield and quality of mulberry, therefore, elucidation of its main pathogenic groups is essential in improving our understanding of this disease and for the development of its potential control measures., Methods: In this study, combined metagenomic sequencing and culture-dependent approaches were used to investigate the microbiome of healthy and bacterial wilt mulberry samples., Results: The results showed that the healthy samples had higher bacterial diversity compared to the diseased samples. Meanwhile, the proportion of opportunistic pathogenic and drug-resistant bacterial flora represented by Acinetobacter in the diseased samples was increased, while the proportion of beneficial bacterial flora represented by Proteobacteria was decreased. Ralstonia solanacearum species complex (RSSC), Enterobacter cloacae complex (ECC), Klebsiella pneumoniae , K. quasipneumoniae , K. michiganensis , K. oxytoca , and P. ananatis emerged as the main pathogens of the mulberry bacterial wilt., Discussion: In conclusion, this study provides a valuable reference for further focused research on the bacterial wilt of mulberry and other plants., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Yuan, Qazi, Li, Yang, Yang, Zhang, Liu and Liu.)

Ralstonia solanacearum, a species complex of bacterial plant pathogens divided into four monophyletic phylotypes, causes plant diseases in tropical climates around the world. Some strains exhibit a broad host range on solanaceous hosts, while others are highly host-specific as for example some banana-pathogenic strains.Previous studies showed that transcription activator-like (TAL) effectors from Ralstonia, termed RipTALs, are capable of activating reporter genes in planta, if these are preceded by a matching effector binding element (EBE). RipTALs target DNA via their central repeat domain, where one repeat pairs with one DNA-base of the given EBE. The repeat variable diresidue dictates base repeat specificity in a predictable fashion, known as the TALE code.In this work, we analyze RipTALs across all phylotypes of the Ralstonia solanacearum species complex. We find that RipTALs are prevalent in phylotypes I and IV but absent from most phylotype III and II strains (10/12, 8/14, 1/24 and 1/5 strains contained a RipTAL, respectively).RipTALs originating from strains of the same phylotype show high levels of sequence similarity (>98%) in the N-terminal and C-terminal regions, while RipTALs isolated from different phylotypes show 47-91% sequence similarity in those regions, giving rise to four RipTAL classes. We show that, despite sequence divergence, the base preference for guanine, mediated by the N-terminal region, is conserved across RipTALs of all classes.Using the number and order of repeats found in the central repeat domain, we functionally sub-classify RipTALs, introduce a new simple nomenclature, and predict matching EBEs for all seven distinct RipTALs identified. We experimentally study RipTAL EBEs and uncover that some RipTALs are able to target the EBEs of other RipTALs, referred to as cross-reactivity. In particular, RipTALs from strains with a broad host range on solanaceous hosts cross-react on each other’s EBEs.Investigation of sequence divergence between RipTAL repeats allows for a reconstruction of repeat array biogenesis, for example through slipped strand mispairing or gene conversion. Using these studies we show how RipTALs of broad host range strains evolved convergently towards a shared target sequence. Finally we discuss the differences between TALE-likes of plant pathogens in the context of disease

Bacterial wilt caused by Ralstonia solanacearum ranks the second top important bacterial plant disease worldwide. It is also the most important bacterial disease threatening the healthy development of Casuarina equisetifolia protection forest. 3-hydroxypalmitic acid methyl ester (3-OH PAME) functions as an important quorum sensing (QS) signal regulating the expression of virulence genes in R. solanacearum , and has been regarded as an ideal target for disease prevention and control. To screen native microorganisms capable of degrading 3-OH PAME, samples of C. equisetifolia branches and forest soil were collected and cultured in the medium containing 3-OH PAME as the sole carbon source. Bacteria with over 85% degradation rates of 3-OH PAME after 7-day incubation were further separated and purified. As a result, strain Q1-7 isolated from forest soil and strain Q4-3 isolated from C. equisetifolia branches were obtained and identified as Pseudomonas novel species Pseudomonas forestsoilum sp. nov. and P. tohonis , respectively, according to whole genome sequencing results. The degradation efficiencies of 3-OH PAME of strains Q1-7 and Q4-3 were 95.80% and 100.00% at 48 h, respectively. Both strains showed high esterase activities and inhibited R. solanacearum exopolysaccharide (EPS) and cellulase production. Application of strains Q1-7 and Q4-3 effectively protects C. equisetifolia , peanut and tomato plants from infection by R. solanacearum . Findings in this study provide potential resources for the prevention and control of bacterial wilt caused by R. solanacearum , as well as valuable materials for the identification of downstream quenching genes and the research and development of quenching enzymes for disease control., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Wang, Hu, Chen, Li, Xue, Song, Qi, Liu, Zhou, Zhang and Zhou.)

Background: Tobacco bacterial wilt (TBW) and black shank (TBS) are responsible for substantial economic losses worldwide; however, microbial interactions and metabolisms in response to TBW and TBS pathogens in the tobacco rhizosphere remain unclear., Methods: We explored and compared the response of rhizosphere microbial communities to these two plant diseases with the incidences in moderate and heavy degrees by sequencing of 16S rRNA gene amplicons and bioinformatics analysis., Results and Discussions: We found that the structure of rhizosphere soil bacterial communities was significantly ( p < 0.05) changed from the incidences of TBW and TBS, which also led to decreased Shannon diversity and Pielou evenness. Compared with the healthy group (CK), the OTUs with significantly ( p < 0.05) decreased relative abundances were mostly affiliated with Actinobacteria (e.g., Streptomyces and Arthrobacter ) in the diseased groups, and the OTUs with significantly ( p < 0.05) increased relative abundances were mainly identified as Proteobacteria and Acidobacteria. Also, molecular ecological network analysis showed that the nodes (<467) and links (<641) were decreased in the diseased groups compared with the control group (572; 1056), suggesting that both TBW and TBS weakened bacterial interactions. In addition, the predictive functional analysis indicated that the relative abundance of genes related to the biosynthesis of antibiotics (e.g., ansamycins and streptomycin) was significantly ( p < 0.05) decreased due to incidences of TBW and TBS, and antimicrobial tests showed that some Actinobacteria strains (e.g., Streptomyces ) and their secreted antibiotics (e.g., streptomycin) could effectively inhibit the growth of these two pathogens., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Tang, Liu, Teng, Xiao, Cai, Wang, Xiao and Chen.)

Ralstonia solanacearum, a species complex of bacterial plant pathogens divided into four monophyletic phylotypes, causes plant diseases in tropical climates around the world. Some strains exhibit a broad host range on solanaceous hosts, while others are highly host-specific as for example some banana-pathogenic strains. Previous studies showed that transcription activator-like (TAL) effectors from Ralstonia, termed RipTALs, are capable of activating reporter genes in planta, if these are preceded by a matching effector binding element (EBE). RipTALs target DNA via their central repeat domain (CRD), where one repeat pairs with one DNA-base of the given EBE. The repeat variable diresidue dictates base repeat specificity in a predictable fashion, known as the TALE code. In this work, we analyze RipTALs across all phylotypes of the Ralstonia solanacearum species complex. We find that RipTALs are prevalent in phylotypes I and IV but absent from most phylotype III and II strains (10/12, 8/14, 1/24, and 1/5 strains contained a RipTAL, respectively). RipTALs originating from strains of the same phylotype show high levels of sequence similarity (>98%) in the N-terminal and C-terminal regions, while RipTALs isolated from different phylotypes show 47–91% sequence similarity in those regions, giving rise to four RipTAL classes. We show that, despite sequence divergence, the base preference for guanine, mediated by the N-terminal region, is conserved across RipTALs of all classes. Using the number and order of repeats found in the CRD, we functionally sub-classify RipTALs, introduce a new simple nomenclature, and predict matching EBEs for all seven distinct RipTALs identified. We experimentally study RipTAL EBEs and uncover that some RipTALs are able to target the EBEs of other RipTALs, referred to as cross-reactivity. In particular, RipTALs from strains with a broad host range on solanaceous hosts cross-react on each other’s EBEs. Investigation of sequence divergence between RipTAL repeats allows for a reconstruction of repeat array biogenesis, for example through slipped strand mispairing or gene conversion. Using these studies we show how RipTALs of broad host range strains evolved convergently toward a shared target sequence. Finally, we discuss the differences between TALE-likes of plant pathogens in the context of disease ecology. [ABSTRACT FROM AUTHOR]

As one of the most important and largest transcription factors, WRKY plays a critical role in plant disease resistance. However, little is known regarding the functions of the WRKY family in cultivated peanuts ( Arachis hypogaea L.). In this study, a total of 174 WRKY genes ( AhWRKY ) were identified from the genome of cultivated peanuts. Phylogenetic analysis revealed that AhWRKY proteins could be divided into four groups, including 35 (20.12%) in group I, 107 (61.49%) in group II, 31 (17.82%) in group III, and 1 (0.57%) in group IV. This division is further supported by the conserved motif compositions and intron/exon structures. All AhWRKY genes were unevenly located on all 20 chromosomes, among which 132 pairs of fragment duplication and seven pairs of tandem duplications existed. Eighteen miRNAs were found to be targeting 50 AhWRKY genes. Most AhWRKY genes from some groups showed tissue-specific expression. AhWRKY46, AhWRKY94, AhWRKY156, AhWRKY68, AhWRKY41, AhWRKY128, AhWRKY104, AhWRKY19, AhWRKY62, AhWRKY155, AhWRKY170, AhWRKY78, AhWRKY34, AhWRKY12, AhWRKY95 , and AhWRKY76 were upregulated in ganhua18 and kainong313 genotypes after Ralstonia solanacearum infection. Ten AhWRKY genes ( AhWRKY34, AhWRKY76, AhWRKY78, AhWRKY120, AhWRKY153, AhWRKY155, AhWRKY159, AhWRKY160, AhWRKY161 , and AhWRKY162 ) from group III displayed different expression patterns in R. solanacearum sensitive and resistant peanut genotypes infected with the R. solanacearum . Two AhWRKY genes ( AhWRKY76 and AhWRKY77 ) from group III obtained the LRR domain. AhWRKY77 downregulated in both genotypes; AhWRKY76 showed lower-higher expression in ganhua18 and higher expression in kainong313. Both AhWRKY76 and AhWRKY77 are targeted by ahy-miR3512, which may have an important function in peanut disease resistance. This study identified candidate WRKY genes with possible roles in peanut resistance against R. solanacearum infection. These findings not only contribute to our understanding of the novel role of WRKY family genes but also provide valuable information for disease resistance in A. hypogaea ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yan, Jin, Raza, Huang, Gu and Zou.)