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Bacterial wilt disease caused by Ralstonia solanacearum (Smith) Yabuuchi is one of the most important bacterial diseases in the world. This study aimed to test some chemical compounds such as bactericides that were applied to control bacterial wilt disease under artificial inoculation conditions and their effects on the yield of potatoes under greenhouse conditions.In vitro, tested chemical compounds (Cefalexin, Gentamycin, and Copper sulphate) inhibit the growth of R. solanacerum compared with control treatment. Copper sulphate was the most effective where the Inhibition zone diameter was 6.0 to7.4mm compared with cefalexin and gentamycin, 3.6 to 5.8mm respectively, but cefalexin was moderately effective 4.2 to 5.8mm and gentamycin was less effective 3.6 to 5.8mm against the pathogen. Inhibition zone diameter was increased with increasing concentrations of tested chemical compounds. Greenhouse experiments showed that the chemical compounds used reduced the severity of potato bacterial wilt disease and increased potato yield compared with control treatment. When using Cefalexin, the mean of wilted shoots was 20.5 and the severity of disease 16.3, while when using Gentamycin as an Abiotic, the mean of wilted shoots was 22.8 and severity of disease 17.6 which was the most effective compared with Copper sulphate treatment were wilted shoot was 31.8 and severity of disease 29.6. The application of tested chemical compounds as soil drench treatment led to a percentage of infection which ranged from16.4 to19.0 more effective than tuber treatment where the percentage of infection ranged from 18.4 to 20.1.

Wilt caused by the bacteria Ralstonia solanacearum, Enterobacter cloacae and Burkholderia glumae, independently or in complex, in Solanaceous vegetables is leading to enormous crop losses. The resistance mechanisms in tomato, chilli peppers, eggplant and potato are polygenically governed and the molecular level analyses have been successful to understand the key genes and pathways involved. QTL and candidate gene mapping, development of molecular markers explaining significant level of phenotypic variation, transcriptome analysis, expression analysis for the putative candidate genes and metabolome analyses which have been successful to reveal the genes and pathways in resistance are discussed in this paper. Genetic transformations using the key genes thus identified have been successful to generate the resistant lines. In addition to a systematic discussion on these topics, the major QTLs for bacterial wilt resistance in tomato and chilli peppers have been analyzed, contributing genes identified and annotated and the mechanisms conserved between these crops are discussed.

Abstract Background Bacteriophages are effective biocontrol strategy as well as ecofriendly remedy for the emerging antibiotic and chemical resistance in bacterial phytopathogens such as bacterial wilt-causing Ralstonia solanacearum. One of the major challenges in the use of bacteriophage therapy for agricultural phytopathogens is maintaining their viability even during variations in pH, temperature, ultraviolet irradiation, and desiccation during field application for sustainable agriculture. Results In this study, the isolation and characterization of phage ɸsp1 for its efficacy against wilt-causing R. solanacearum performed on Solanum lycopersicum (tomato) seedlings and Solanum tuberosum (potato) tuber assay are reported. Bacteriophage was found to be viable and stable at a wide pH range (3.0–9.0) and at temperatures up to 55 °C. Phage ɸsp1 required ~15 min for adsorption and completed its life cycle in 25–30 min by host cell lysis with a burst size of ~250–300. Phage ɸsp1 eradicated 94.73% preformed R. solanacearum biofilm and inhibited biofilm formation by 73.68% as determined by the static crystal violet microtiter biofilm assay. Transmission electron microscope revealed the phage ɸsp1 to be approximately 208±15 nm in size, comprising of icosahedral head (100 ±15 nm) and tail, as belonging to Myoviridae family. Plant bioassays showed 81.39 and 87.75% reduction in pathogen count using phages ɸsp1 in potato tuber and tomato seedlings, respectively. Reversal in disease symptoms was 100% in phage-treated tuber and tomato plant (pot assay) compared to only pathogen-treated controls. Conclusion Isolated bacteriophage ɸsp1 was found to be highly host specific, effective in biofilm prevention, and capable of inhibiting bacterial wilt at low multiplicity of infection (1.0 MOI) in tomato as well as potato tuber bioassays. Phages ɸsp1 were environmentally stable as they survive at variable pH and temperature. Bacteriophage ɸsp1 shows a promise for development into a biocontrol formulation for the prevention of R. solanacearum bacterial wilt disease.

The Ralstonia solanacearum species complex causes bacterial wilt in a variety of crops. Tomato cultivar Hawaii 7996 is a widely used resistance resource; however, the resistance is evaded by virulent strains, with the underlying mechanisms still unknown. Here, we report that the phylotype Ⅱ strain ES5-1 can overcome Hawaii 7996 resistance. RipV2, a type Ⅲ effector specific to phylotype Ⅱ strains, is vital in overcoming tomato resistance. RipV2, which encodes an E3 ubiquitin ligase, suppresses immune responses and Toll/interleukin-1 receptor/resistance nucleotide-binding/leucine-rich repeat (NLR) (TNL)-mediated cell death. Tomato helper NLR N requirement gene 1 (NRG1), enhanced disease susceptibility 1 (EDS1), and senescence-associated gene 101b (SAG101b) are identified as RipV2 target proteins. RipV2 is essential for ES5-1 virulence in Hawaii 7996 but not in SlNRG1 -silenced tomato, demonstrating SlNRG1 to be an RipV2 virulence target. Our results dissect the mechanisms of RipV2 in disrupting immunity and highlight the importance of converged immune components in conferring bacterial wilt resistance. [Display omitted] • Ralstonia solanacearum phylotype Ⅱ strain ES5-1 overcomes the resistance of Hawaii 7996 • Type Ⅲ effector RipV2 contributes to ES5-1 virulence in Hawaii 7996 • RipV2 ubiquitinates and degrades tomato SlNRG1-SlEDS1-SlSAG101b immune complex • RipV2 inhibits SlNRG1-mediated immune responses, cell death, and bacterial wilt resistance Qi et al. reveal that RipV2, a type Ⅲ effector specifically present in Ralstonia solanacearum phylotype Ⅱ strains, ubiquitinates and degrades tomato SlNRG1, SlEDS1, and SlSAG101b to dampen Hawaii 7996 resistance. This finding elucidates the mechanism by which bacterial pathogen counteracts host defense. [ABSTRACT FROM AUTHOR]

Bacterial wilt caused by the β-proteobacterium Ralstonia solanacearum is one of the most destructive soil-borne pathogens in peppers (Capsicum annuum L.) worldwide. Cultivated pepper fields in Korea face a continuous spread of this pathogen due to global warming. The most efficient and sustainable strategy for controlling bacterial wilt is to develop resistant pepper varieties. Resistance, which is quantitatively inherited, occurs differentially depending on R. solanacearum isolates. Therefore, in this study, we aimed to identify resistance quantitative trait loci (QTLs) in two F2 populations derived from self-pollination of a highly resistant pepper cultivar 'Konesian hot' using a moderately pathogenic 'HS' isolate and a highly pathogenic 'HWA' isolate of R. solanacearum for inoculation, via genotyping-by-sequencing analysis. QTL analysis revealed five QTLs, Bwr6w-7.2, Bwr6w-8.1, Bwr6w-9.1, Bwr6w-9.2, and Bwr6w-10.1, conferring resistance to the 'HS' isolate with R2 values of 13.05, 12.67, 15.07, 10.46, and 9.69%, respectively, and three QTLs, Bwr6w-5.1, Bwr6w-6.1, and Bwr6w-7.1, resistant to the 'HWA' isolate with phenotypic variances of 19.67, 16.50, and 12.56%, respectively. Additionally, six high-resolution melting (HRM) markers closely linked to the QTLs were developed. In all the markers, the mean disease index of the paternal genotype was significantly lower than that of the maternal genotype. The QTLs and HRM markers are expected to be useful for the development of pepper varieties with high resistance to bacterial wilt. [ABSTRACT FROM AUTHOR]

The focus of this research was to determine how secondary metabolites from two isolates of Pseudomonas fluorescens (P8 and P60) affected potato plant resistance to bacterial wilt (Ralstonia solanacearum) as well as potato growth and productivity. For four months, this experiment was carried out at Serang Village's potato field, in Karangreja District, Purbalingga Regency, at an altitude of 1285 m above sea level, with Andisol soils, an average temperature of 22.31°C, and relative humidity of 84.09%. A randomized block design was used with a control treatment, drenching with 1.5 g/L bactericide (20% streptomicin sulphate) administered six times, and drenching with secondary metabolites of P. fluorescens P8 or P. fluorescens P60 administered three, six, nine, and 12 times. Each treatment was carried out five times. Each treatment resulted in a different incubation period, disease intensity, infection rate, plant height, number of tubers, tuber weight per plant, wet and dry weight of crop, fresh and dry weight of root, number of branches, and phenolic compound analysis. The results showed that applying P. fluorescens P8 and P. fluorescens P60 secondary metabolites to potato plants can induce resistance by increasing the content of phenolic compounds in the plants. Drenching with secondary metabolites from P. fluorescens P8 or P. fluorescens P60 12 times can reduce the incubation period by 9.23%, the intensity of disease by 75%, and the incidence of disease by 53.57%. Plant height, crop dry weight, root fresh weight, root dry weight, number of tubers, and tuber weight per plant can all be increased by using secondary metabolites. [ABSTRACT FROM AUTHOR]

Bacterial wilt disease caused by Ralstonia solanacearum impacts negatively on potato production. This study evaluated the effects of thymol and eugenol encapsulated in chitosan nanoparticles (TCNP and ECNP) on the virulence genes (PhcA, XpsR and HrpG) of R. solanacearum and their in vivo efficacy against bacterial wilt. Gene expression levels of virulence genes in the presence of TCNP (5.6 and 11.3 µg mL−1) and ECNP (11.3 and 22.5 µg mL−1) together with those of plant defense genes (chitinase and β-l,3-glucanase) were determined through RT-qPCR. All the virulence genes were downregulated when exposed to both TCNP and ECNP while glucanase and chitinase genes increased and peaked after 18 hours post inoculation. The lowest disease severity index (10.3%) was recorded with plants treated with 90 µg mL−1 ECNP. The results of this study show that both TCNP and ECNP have a potential to be used as bacterial wilt bactericides. [ABSTRACT FROM AUTHOR]

The germicidal efficacy of various physico-chemical agents against culturable bacterial cells and bacterial spores is well studied and reviewed; except for the dormant Viable But Non Culturable state (VBNCs). VBNC is a special physiological state, where the bacterium is not culturable in culture media, but remains alive, infective and virulent. The aim of this study is to evaluate the germicidal efficacy of three physico-chemical agents against the VBNC forms of destructive phytopathogen Ralstonia pseudosolanacearum. Effect of thermotherapy Formalin and Sodium Hypochlorite on these forms was evaluated using standard plate count and reverse culturability tests. The disinfectants were found equally effective against the culturable cells and VBNC forms at recommended concentrations. [ABSTRACT FROM AUTHOR]

Summary: Tomato varieties resistant to the bacterial wilt pathogen Ralstonia solanacearum have the ability to restrict bacterial movement in the plant. Inducible vascular cell wall reinforcements seem to play a key role in confining R. solanacearum into the xylem vasculature of resistant tomato. However, the type of compounds involved in such vascular physico‐chemical barriers remain understudied, while being a key component of resistance.Here we use a combination of histological and live‐imaging techniques, together with spectroscopy and gene expression analysis to understand the nature of R. solanacearum‐induced formation of vascular coatings in resistant tomato.We describe that resistant tomato specifically responds to infection by assembling a vascular structural barrier formed by a ligno‐suberin coating and tyramine‐derived hydroxycinnamic acid amides. Further, we show that overexpressing genes of the ligno‐suberin pathway in a commercial susceptible variety of tomato restricts R. solanacearum movement inside the plant and slows disease progression, enhancing resistance to the pathogen.We propose that the induced barrier in resistant plants does not only restrict the movement of the pathogen, but may also prevent cell wall degradation by the pathogen and confer anti‐microbial properties, effectively contributing to resistance. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum is a causative agent of bacterial wilt and therefore poses a serious threat to cultivated peppers (Capsicum annuum L.). Although attempts have been made to control bacterial wilt by grafting, the disease resistance mechanisms that protect grafted peppers are poorly understood. Here, we grew grafted peppers composed of the rootstock Buyeding or Weishi and the scion Xinfeng 2. Following infection by R. solanacearum, we assessed the differences in lipid peroxidation, cellular structure, root secondary metabolism, and biomass, between grafted plants and controls. The grafted plants exhibited a greater root biomass than the control plants after infection. The root cell ultrastructure of the grafted plants showed only slight injury relative to that in the controls, and the roots of the grafted peppers were partially resistant to R. solanacearum. Grafted pepper plants showed lower levels of lipid peroxidation. Lignin content, salicylic acid levels, and the activities of phenylalanine ammonia lyase (PAL), peroxidase (POD), catalase (CAT), and polyphenol oxidase (PPO), were also higher in grafted plants. All of these effects occurred concomitantly with increased R. solanacearum resistance. Taken together, our findings demonstrate that grafting can significantly improve the disease resistance of pepper. Moreover, our results suggest that the Weishi rootstock may be very useful for the prevention and control of bacterial wilt in cultivated peppers. [ABSTRACT FROM AUTHOR]

Ginger is one of the most widely cultivated spice grown in various cropping systems and locations throughout the southwestern Ethiopia. Bacterial wilt, caused by Ralstonia solanacearum, is one of the serious diseases of ginger in Ethiopia. Field experiments were conducted during 2019 and 2020 to assess effects of soil amendments on bacterial wilt development and epidemics at Tepi, southwestern Ethiopia. Three soil amendments practices: compost, effective microorganisms and Bordeaux mixture alone and in integration were evaluated. Treatments were arranged in a randomized complete block design with three replications. Compost at the rate of 7 ton/ha enriched with effective microorganisms and Bordeaux mixture treatment significantly reduced ginger bacterial wilt incidence, area under disease progress curve and disease progress rate. This treatment reduced bacterial wilt mean incidence up to 21.08 % as compared to untreated control plot. Compost at the rate of 7 ton/ha also slowed down epidemic progression of bacterial wilt and significantly reduced the disease parameters when effective microorganisms and Bordeaux mixture were integrated. The overall results indicated that integrated compost enriched with effective microorganisms and Bordeaux mixture was effective to slow down the epidemics of ginger bacterial wilt and sustain ginger production and productivity. Hence, integrated compost enriched with effective microorganisms and Bordeaux mixture along with other crop management systems are recommended to improve ginger production and productivity at southwestern Ethiopia and other similar ecologies. [ABSTRACT FROM AUTHOR]

Every year, despite the use of chemicals, significant crop loss is caused by pathogenic microbes. Plant innate resistance to pathogens depends on two sets of genetically encoded immune receptors that sense invaders and trigger signaling cascades leading to reinforcement of physical barriers and production of various antimicrobial compounds. In the past 30 years, the molecular cloning and characterization of plant immune receptors have deepened our understanding of the plant immune system and more importantly, have provided means to improve crop protection against devastating pathogens. Here, we review the molecular characterization of selected immune receptors that can detect multiple species of bacterial pathogens through an expanded recognition range, or through the detection of conserved pathogen activities or host targets. These recent structural and molecular insights about the activation of immune receptors provide the necessary framework to design their concomitant deployment in crops, in order to lower selective pressure on pathogen populations and prevent evasion from recognition. Hence, these few immune receptors emerge as high potential genetic resources to provide durable and environmentally safe protection against important bacterial diseases of solanaceous crops. [ABSTRACT FROM AUTHOR]

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Survival niches for phytopathogenic bacteria are important for disease management. We study the survival of Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff), the causal agent of bacterial wilt in common bean and bacterial tan spot in soybean, in the soil. While Cff strains from common bean already have several studies, the ecology of soybean strains needs more information. We used the strain Feij.2628A from common bean, and CffR1, Cff2R, and Cff4R from soybean. The soils were infested with bacterial suspension (107 CFU.mL−1) from each of the strains. For experiments under laboratory conditions, they were incubated in BOD incubator (biochemical oxygen demand) at 20 °C. Under natural field conditions, pots were maintained in the field without supplementary irrigation. The survival of Cff strains was evaluated every seven days until they were not detected. The identity of the bacterium was confirmed in all treatments by PCR. Under laboratory conditions, the survival period of Feij.2628A ranged from 140 to 154 days, and between 77 to 119 days for the soybean strains. Feij.2628A and Cff1R survived for a maximum of 91 days in soil with higher clay content, under natural field conditions. Cff survival was negatively influenced by high temperatures, low soil moisture, and precipitation in some treatments. These results demonstrated the potential survival of Cff strains from common bean and soybean in the soil, being potential inoculum sources for subsequent cultivations of these crops. Fallow periods ranging from three to four months may reduce the amount of inoculum in soils under these conditions. [ABSTRACT FROM AUTHOR]

Bacterial wilt caused by Ralstonia solanacearum is one of the most important bacterial diseases in potato production. This study aimed to obtain the transgenic potato (Solanum tuberosum L.) cultivar IPB CP3, containing LYZ‐C gene encoding for lysozyme type C, resistant to bacterial disease caused by R. solanacearum. Genetic transformation using Agrobacterium tumefaciens LBA4404 to 124 internode explants resulted in the transformation efficiency of about 47.58% with a regeneration efficiency of approximately 30.51%. Gene integration analysis showed that 16 clones were confirmed as transgenic clones containing the LYZ‐C gene. Analysis of resistance to R. solanacearum of three transgenic clones showed that all three transgenic clones were more resistant than a non‐transgenic one. This result showed that the LYZ‐C gene integrated in the genome of transgenic potato increased the resistance of potato plants to R. solanacearum. We obtained two transgenic clones considered resistant to bacterial wilt disease.

Abstract Background Ralstonia solanacearum is the causal agent of bacterial wilt, a devastating plant disease responsible for serious economic losses especially on potato, tomato, and other solanaceous plant species in temperate countries. In R. solanacearum, gene expression analysis has been key to unravel many virulence determinants as well as their regulatory networks. However, most of these assays have been performed using either bacteria grown in minimal medium or in planta, after symptom onset, which occurs at late stages of colonization. Thus, little is known about the genetic program that coordinates virulence gene expression and metabolic adaptation along the different stages of plant infection by R. solanacearum. Results We performed an RNA-sequencing analysis of the transcriptome of bacteria recovered from potato apoplast and from the xylem of asymptomatic or wilted potato plants, which correspond to three different conditions (Apoplast, Early and Late xylem). Our results show dynamic expression of metabolism-controlling genes and virulence factors during parasitic growth inside the plant. Flagellar motility genes were especially up-regulated in the apoplast and twitching motility genes showed a more sustained expression in planta regardless of the condition. Xylem-induced genes included virulence genes, such as the type III secretion system (T3SS) and most of its related effectors and nitrogen utilisation genes. The upstream regulators of the T3SS were exclusively up-regulated in the apoplast, preceding the induction of their downstream targets. Finally, a large subset of genes involved in central metabolism was exclusively down-regulated in the xylem at late infection stages. Conclusions This is the first report describing R. solanacearum dynamic transcriptional changes within the plant during infection. Our data define four main genetic programmes that define gene pathogen physiology during plant colonisation. The described expression of virulence genes, which might reflect bacterial states in different infection stages, provides key information on the R. solanacearum potato infection process.

Ginger (Zingiber officinale Rosc.) is a type of rhizome plant that has high potential to be developed in Indonesia as a medicinal plant or spice. Inhibiting factors in ginger production is Ralstonia solanacearum which causes bacterial wilt disease. Rhizobacteria is a potential biological agent to control the disease. It is necessary to review from various study on rhizobacteria both the use of antagonistic bacteria singly or in a consortia. Aimed to analyze various sources of scientific literature regarding the potential of antagonistic bacteria singly or consortia compared to synthetic bactericides in controlling of R. solanacearum. The results showed that three recommended antagonist bacteria were found, namely Pseudomonas fluorescens, Bacillus subtilis, and Bacillus thuringiensis. which has the potential to control bacterial wilt disease. The rhizobacterial consortia are more effective as a sustainable, safe and environmentally friendly control on bacterial wilt disease with a percentage of disease incidence 10%. In addition, the rhizobacterial consortia are three and six times more effective than the single antagonistic bacteria and bactericides respectively.

Abstract Background Ralstonia solanacearum causes bacterial wilt disease in tomato and other crops resulting in huge economic losses worldwide. Several measures have been explored for the control of R. solanacearum, but the desired control level of the disease through sustainable and ecofriendly way is still awaited. Main body In this study, fungal metabolites produced by Trichoderma harzianum were investigated in the form of crude extract for the management of R. solanacearum both in vitro and in planta in tomato plants. In in vitro investigation, fungal metabolites were checked for their antibacterial potential at different concentrations (30, 60, 90, 120, 150, and 180 mg ml−1) and bacterial cell morphology was observed under scanning electron microscopy (SEM). In a greenhouse experiment, different application times (0, 3, and 6 days before transplantation DBT) and doses (0, 3, 6, and 9%) of the fungal metabolites were tested for their effects on soil bacterial population, disease severity and plant growth of tomato plants. The in vitro evaluation showed a strong antibacterial activity of fungal metabolites in concentration dependent manner. The highest concentration 180 mg ml−1 produced maximum inhibition zone (20.2 mm) having non-significant difference with the inhibition zone (20.5 mm) produced by the standard antibiotic streptomycin. The SEM analysis revealed severe morphological destructions of bacterial cells. In case of greenhouse experiment, the highest decrease in soil bacterial population, lowest disease severity, and maximum increase in plant growth parameters were obtained by highest dose (9%) and longest application time (6 DBT). Conclusion The fungal metabolites produced by T. harzianum could be used as low-cost, environment-friendly, and sustainable management strategy for the control of R. solanacearum in tomato plants.

Common bean (Phaseolus vulgaris) is one of the major legume crops cultivated worldwide. Bacterial wilt (BW) of common bean (Curtobacterium flaccumfaciens pv. flaccumfaciens), being a seed-borne disease, has been a challenge in common bean producing regions. A genome-wide association study (GWAS) was conducted to identify SNP markers associated with BW resistance in the USDA common bean core collection. A total of 168 accessions were evaluated for resistance against three different isolates of BW. Our study identified a total of 14 single nucleotide polymorphism (SNP) markers associated with the resistance to BW isolates 528, 557, and 597 using mixed linear models (MLMs) in BLINK, FarmCPU, GAPIT, and TASSEL 5. These SNPs were located on chromosomes Phaseolus vulgaris [Pv]02, Pv04, Pv08, and Pv09 for isolate 528; Pv07, Pv10, and Pv11 for isolate 557; and Pv04, Pv08, and Pv10 for isolate 597. The genomic prediction accuracy was assessed by utilizing seven GP models with 1) all the 4,568 SNPs and 2) the 14 SNP markers. The overall prediction accuracy (PA) ranged from 0.30 to 0.56 for resistance against the three BW isolates. A total of 14 candidate genes were discovered for BW resistance located on chromosomes Pv02, Pv04, Pv07, Pv08, and Pv09. This study revealed vital information for developing genetic resistance against the BW pathogen in common bean. Accordingly, the identified SNP markers and candidate genes can be utilized in common bean molecular breeding programs to develop novel resistant cultivars.

ABSTRACT Bacterial wilt is accompanied by microbial communities shift and soil acidification. However, the relationship between the changes of bacterial communities and bacterial wilt under the influence of different acidification levels has not been fully elucidated. Here, we analyzed the abundance of Ralstonia solanacearum, rhizosphere bacterial communities and carbon metabolism at differently acidic levels (pH 6.45, pH 5.60, pH 5.35, pH 4.90 and pH 4.45) and soil amendment treatment (CaO). The results indicated that both the abundance of R. solanacearum and the incidence of bacterial wilt showed a significant trend of first increasing and then decreasing with the increase of soil pH. The Firmicutes phylum and potentially beneficial genera Bacillus, Paenibacillus, Flavobacterium and Pseudomonas were significantly enriched at pH 6.45. The metabolic ability in response to the l-arginine and 4-hydroxybenzoic acid was significantly increased at pH 6.45. After using CaO to increase the pH of diseased soil from 5.45 to 6.05, the abundance of R. solanacearum and the incidence of bacterial wilt were significantly reduced, the Firmicutes and potentially beneficial genera Bacillus and Pseudomonas were significantly enriched. Overall, alleviating soil acidification to a slightly acidic level (pH 6.0–6.5) could suppress bacterial wilt by suppressing the growth of R. solanacearum and enriching the rhizosphere potentially beneficial bacteria, and further emphasized the importance of increasing soil pH in biological control of bacterial wilt. IMPORTANCE The rhizosphere microbiota and soil acidification have been shown to have impacts on bacterial wilt. However, the influence of different acidification levels on the rhizosphere communities and bacterial wilt has not been fully studied. In this study, the potentially beneficial bacteria (Bacillus and Pseudomonas) were significantly enriched in the slightly acidic soil (pH 6.45), leading to the increase of the metabolism of 4-hydroxybenzoic acid and the decrease of pathogenic R. solanacearum, thereby alleviating the occurrence of bacterial wilt. The changes of potentially beneficial bacteria and pathogenic R. solanacearum in strongly acidic soil (pH 5.35) with the highest incidence of bacterial wilt were just the opposite. These findings help clarify the mechanisms by which soil bacteria exert influence on bacterial wilt outbreak under different soil acidification levels.

Ralstonia solanacaerum is one of the most devastating bacteria causing bacterial wilt disease in more than 200 species of plants, especially those belonging to the family Solanaceae. To cope with this pathogen, plants have evolved different resistance mechanisms depending on signal transduction after perception. Phosphorylation is the central regulatory component of the signal transduction pathway. We investigated a comparative phosphoproteomics analysis of the stems of resistant and susceptible tomatoes at 15 min and 30 min after inoculation with Ralstonia solanacearum to determine the phosphorylated proteins involved in induced resistance. Phosphoprotein profiling analyses led to the identification of 969 phosphoproteins classified into 10 functional categories. Among these, six phosphoproteins were uniquely identified in resistant plants including cinnamyl alcohol dehydrogenase 1 (CAD1), mitogen-activated protein kinase kinase kinase 18 (MAPKKK18), phospholipase D delta (PLDDELTA), nicotinamide adenine dinucleotide transporter 1 (NDT1), B3 domain-containing transcription factor VRN1, and disease resistance protein RPM1 (RPM1). These proteins are typically involved in defense mechanisms across different plant species. qRT-PCR analyses were performed to evaluate the level of expression of these genes in resistant and susceptible tomatoes. This study provides useful data, leading to an understanding of the early defense mechanisms of tomatoes against R. solanacearum. [ABSTRACT FROM AUTHOR]

Ralstonia solanacearum species complex is a group of vascular soil‐borne plant pathogens that causes bacterial wilt on various host plants and has caused great yield losses in Thailand. In 2020 and 2021, bacterial wilt diseases were observed in Chiang Mai and Chiang Rai Province, Thailand. Approximately 19 bacterial wilt strains that were infecting 10 plant species were identified. The different bacterial wilt strains were characterized based on biovar test, phylotypes and DNA sequence of egl gene to determine their sequevar classification. The results of biovar tests by oxidation of disaccharides and hexose alcohols showed the strains belong to biovars 2T, 3 and 4. The partial egl gene sequence analysis of the strains clustered into five type groups that included phylotype IIA sequevar 1 as R. solanacearum and phylotype I, which included 4 sequevars (13, 17, 30 and 47) as R. pseudosolanacearum. The results showed that tomato, Thai eggplant and marigold were susceptible to all bacterial wilt strains. Furthermore, R. pseudosolanacearum biovar 2T phylotype I sequevar 17 was specifically pathogenic to pumpkin and cucumber, but sequevar 13 produced bacterial wilt symptoms on cucumber, and the two sequevars were non‐symptomatic on Patumma and turmeric. R. pseudosolanacearum biovar 4 phylotype I sequevar 30 was specifically pathogenic to Patumma and turmeric. This new report expands the knowledge on genetic analysis and host range diversity of the R. solanacearum species complex in Thailand. [ABSTRACT FROM AUTHOR]

Bacterial wilt is a serious disease of potato (Solanum tuberosum L.) caused by the soil-borne pathogenic bacterium Ralstonia solanacearum. Detecting changes in protein abundance in potato plants in response to R. solanacearum is a pivotal step in uncovering the molecular interactions of plant pathogens. In this study, using the disease-resistant cultivar 'Zhongshu 3', we analyzed protein expression in potato seedlings inoculated with R. solanacearum every 12 h for a total of 72 h using isobaric tags for relative and absolute quantitation-based proteomics. Our results indicate that pathogenesis-related proteins, stressrelated proteins, non-specific lipid transfer proteins, small heat shock proteins, and osmotin-like proteins were up-regulated in response to pathogen infection at different time points. The accumulation of these proteins in response to biotic stress suggests that these proteins play an important role in pathogen resistance. Our findings will provide an important basis for characterizing the role of these proteins in increasing plant resistance to pathogens and in breeding bacterial wilt-resistant plants. [ABSTRACT FROM AUTHOR]

The bacterial wilt is a global destructive plant disease that initiated by the phytopathogenic Ralstonia solanacearum. This study display a novel biofabrication of silica/silver nanocomposite using Fusarium oxysporum-fermented rice husk (RH) under solid state fermentation (SSF). The biofabricated nanocomposite was characterized by XRD, UV–Vis. spectroscopy, DLS, SEM, EDX elemental mapping, and TEM analyses as well as investigated for anti-R. solanacearum activity. Response surface methodology was also processed for optimizing the biofabrication process and improving the anti-bacterial activity of the fabricated nanocomposite. Maximum suppression zone of 29.5 mm against R. solanacearum was reached at optimum RH content of 6.0 g, AgNO3 concentration of 2.50 mM, reaction pH of 6.3, and reaction time of 2 days. The anti-R. solanacearum activity of the fabricated nanocomposite was further improved by exposing the F. oxysporum strain to a gamma irradiation dose of 200 Gy. In conclusion, RH recycling under SSF by F. oxysporum could provide an innovative, facile, non-expensive, and green approach for fabricating SiO2/Ag nanocomposite that could be applied efficiently as an eco-friendly antibacterial agent to combat R. solanacearum in agricultural applications. Moreover, the developed method could serve as a significant platform for the designing of new nanostructures for broad applications. Key points: Biofabricated SiO2/Ag nanocomposite by gamma irradiated-F. oxysporum under SSF. Statistical optimization and complete characterization of SiO2/Ag nanocomposite. Application in agriculture for combating the wilt causing R. solanacearum. [ABSTRACT FROM AUTHOR]

Turmeric (Curcuma longa L. (Zingiberaceae)) is a rich source of medicinally important chemical compounds obtained from both pseudostem (aboveground part) and rhizome (underground part). However, the availability of disease-free elite varieties of the plant is highly restricted. Leaf spot caused by Colletotrichum curcumae Butler & Bisby and bacterial wilt caused by Ralstonia pseudosolanacearum Safni have been reported as major diseases responsible for reduced rhizome productivity in the turmeric plant. Meristem culture is a well-known alternative for production of disease-free mother rhizome. The aim of this investigation was to eradicate bacterial and fungal disease instances through culturing of meristematic tissue pretreated with 1.44 to −2.88 μM GA (gibberellic acid) followed by detection of disease contamination using PCR amplification and chemical assay. Meristem (0.5 to 2.0 mm) was identified as a good material for initiating tissue culture, as it exhibited 100% survival rate and absence of R. pseudosolanacearum and C. curcumae when tested with SPA (sucrose peptone agar), TZC (Kelman's tripheny tetrazolium chloride), and PDA (potato dextrose agar) selective media. Plantlets derived from meristematic tissues of size 2.0 mm were healthy and displayed rapid recovery and uniformity in size. Disease-free status of plantlets (derived from meristem culture) was confirmed by PCR products of specific primers, RP1F/RP1R and RP3F/RP3R for R. pseudosolanacearum and CCactF/CCactR and CChisF/CChisR for C. curcumae. Moreover, shoot elongation using supplemented GA in the culture medium was successfully induced for meristem cutting. All meristem cultures pretreated with GA were confirmed to be contamination-free through TZC, SPA, and PDA media as well as PCR amplification. Based on this study, the 1.0 to 2.0 mm in size of meristem cutting and pretreatment with 1.44 to 2.88 μM GA for shoot elongation were successfully identified in addition to confirming the disease-free status of the plantlets using chemical assay and PCR products. [ABSTRACT FROM AUTHOR]

In order to understand the composition of microbial flora in the stem tissue of tobacco plants mixed occurrence of bacterial wilt and black shank disease. Illumina Miseq high-throughput sequencing technology was used to study the structures and diversities of fungal and bacterial communities in diseased and healthy tobacco stems. The results were as follows:(1)The richness and diversity of the fungal community in the diseased stem tissue were lower than those in the healthy stem tissue, and the richness and diversity of the bacterial community in the diseased stem tissue were higher than those in the healthy stem tissue.(2)Cryptococcus, Alternaria and Fusarium were the dominant fungi in the healthy stem tissue, and the sum of the relative abundances of the three genera were more than 80% of the fungal community.(3) Cryptococcus, Alternaria, Fusarium and unclassified_f_Davidiellaceae were the dominant fungi in the diseased stem tissues.(4) norank_c_Cyanobacteria and Ralstonia were the dominant bacteria in diseased stem tissues. The above results indicate that the mixture of bacterial wilt and black shank disease can significantly change the structures and diversities of fungal and bacterial communities, and can also destroy the stability of microbial community in tobacco stem tissue. [ABSTRACT FROM AUTHOR]

Background: Ralstonia solanacearum, one of the most devastating bacterial plant pathogens, is the causal agent of bacterial wilt. Recently, several studies on resistance to bacterial wilt have been conducted using the Arabidopsis-R. solanacearum system. However, the progress of R. solanacearum infection in Arabidopsis is still unclear. Results: We generated a bioluminescent R. solanacearum by expressing plasmid-based luxCDABE. Expression of luxCDABE did not alter the bacterial growth and pathogenicity. The light intensity of bioluminescent R. solanacearum was linearly related to bacterial concentrations from 104 to 108 CFU·mL−1. After root inoculation with bioluminescent R. solanacearum strain, light signals in tomato and Arabidopsis were found to be transported from roots to stems via the vasculature. Quantification of light intensity from the bioluminescent strain accurately reported the difference in disease resistance between Arabidopsis wild type and resistant mutants. Conclusions: Bioluminescent R. solanacearum strain spatially and quantitatively measured bacterial growth in tomato and Arabidopsis, and offered a tool for the high-throughput study of R. solanacearum-Arabidopsis interaction in the future. [ABSTRACT FROM AUTHOR]

Pogostemon cablin (patchouli), an important medicinal and aromatic plant, is widely used in traditional Chinese medicine as well as in perfume industry. Patchouli plants are susceptible to bacterial wilt disease, which causes significant economic losses by reduction in yield and quality of the plant products. However, few studies focus on the pathogens causing bacterial wilt on patchouli. In this study, strain Pa82 was isolated from diseased patchouli plants with typical bacterial wilt symptoms in Guangdong province, China, and was confirmed to be a highly virulent pathogen of patchouli bacterial wilt. Comparative sequence analysis of 16S rRNA gene showed that the strain was closely related to Kosakonia sp. CCTCC M2018092 (99.9% similarity) and Kosakonia cowanii Esp_Z (99.8% similarity). Moreover, phylogenetic tree based on 16S rRNA gene sequences showed that the strain was affiliated with genus Kosakonia. Further, the whole genome of strain Pa82 was sequenced, and the sequences were assembled and annotated. The complete genome of the strain consists of one chromosome and three plasmids. Average nucleotide identity (ANI) and phylogenetic analysis revealed that the strain belongs to Kosakonia cowanii (designated Kosakonia cowanii Pa82). Virulence-related genes of the strain involved in adherence, biofilm formation, endotoxin and other virulence factors were predicted. Among them, vgrG gene that encodes one of the type VI secretion system components was functionally validated as a virulence factor in Kosakonia cowanii Pa82 through construction of Tn 5 insertion mutants and identification of mutant defective in virulence. [ABSTRACT FROM AUTHOR]

Blueberry production is expanding rapidly in the United States and globally. In 2016, bacterial wilt was discovered in Florida blueberry production. Because of the international movement of plants, this disease poses a significant risk to production. The purpose of this project was to evaluate the resistance of blueberry species and cultivars to the three genetically distinct populations of Ralstonia identified in Florida blueberry production. Nineteen cultivars/species of Vaccinium were used in this study. Plants were selected from wild, northern, southern, midbush, lowbush, and rabbiteye varieties. Plants were inoculated with three genetically distinct strains of the pathogen, and experiments were replicated three times. Varying levels of quantitative (multigenic) bacterial wilt resistance were observed among varieties tested, with rabbiteye cultivars being the most resistant. These results are similar to bacterial wilt resistance observed in other agronomic crops. We also observed a unique specific qualitative (vertical) resistance in the rabbiteye cultivar ´Ochlockonee'. The pathogen was unable to colonize plant tissue and cause disease on this cultivar. This is the first report of qualitative resistance to Ralstonia solanacearum in blueberry. Both the multigenic and vertical resistance can be incorporated into blueberry breeding programs to mitigate potential losses to bacterial wilt. [ABSTRACT FROM AUTHOR]

As tobacco bacterial wilt is a bacterial disease that is increasingly harmful to tobacco plants′ growth, it is of great significance to explore the changes in the soil microbial community structure and its biological regulation mechanism after tobacco plant suffers from bacterial wilt. Field experiments were conducted to investigate the differences of bacterial community structure among four treatments： ZCTR（conventional fertilization, rhizosphere soil of healthy tobacco plants）, ZCSW（application of 1.2 t·hm-2 biochar, rhizosphere soil of healthy tobacco plants）, QKTR（conventional fertilization, rhizosphere soil of bacterial wilt tobacco plants）, and QKSW（application of 1.2 t·hm-2 biochar, rhizosphere soil of bacterial wilt tobacco plants）. After the biochar application, the bacteria′s α diversity increased, albeit without a significant difference. The bacterial community richness and structure changed. Under the same fertilization conditions, the bacterial α diversity and community abundance of bacterial wilt tobacco plants in the rhizosphere soil were less than those of healthy soil. The bacterial α diversity and community richness of the rhizosphere soil of diseased tobacco plants without biochar were higher than that of healthy tobacco plants, the abundance of Gemmatimonadetes and Acidobacteria increased by 0.26 and 1.27 percent points, while the abundance of Proteobacteria and Actinobacteria decreased by 1.08 and 0.14 percent points. Compared with no biochar treatment, the plant height, leaf length, leaf width and stem girth of the biochar application treatment increased by 11.83%, 16.88%, 6.70% and 10.80%, respectively, and the disease index decreased. The incidence rate of the bacterial wilt was reduced by 9.1 percent points with the biochar application treatment compared with the no biochar treatment. Studies have shown that biochar application can alleviate the decline in bacterial abundance caused by bacterial wilt, reduce the relative abundance of pathogenic bacteria, and drive the increase in the relative abundance of functional growth-promoting bacteria（Gemmatimonadetes）. This reduces the incidence of bacterial wilt and preliminarily clarifies the biochar′s mechanism on the micro-ecological regulation of bacterial wilt in tobacco-growing soil. [ABSTRACT FROM AUTHOR]

In order to explore the beneficial microbial resources for disease-suppressive soils against tobacco bacterial wilt, field experiments were conducted to analyze the microbial community structure and the relative abundances of Ralstonia solanacearum and antagonistic bacteria in disease-suppressive soils and disease-conducive soils at different altitudes. The results showed that at phylum level, the relative abundances of beneficial bacteria (Actinobacteria, Firmicutes, Cyanobacteria) and beneficial fungi (Zygomycota, Chytridiomycota) in disease-suppressive soils were higher than those in disease-conducive soils. At genus level, the relative abundances of some beneficial microorganisms (Mesorhizobium, Nitrosospira) and antagonistic microorganisms (Pseudomonas, Flavobacterium, Rhizobium, Aspergillus, Trichoderma, Myrothecium) in disease-suppressive soils were significantly higher than those in disease-conducive soils, which all had obviously negative correlations with the incidence of bacterial wilt. However, the relative abundance of pathogenic microorganism (Ralstonia) in disease-suppressive soils was significantly lower than that in disease-conducive soils. To sum up, there were abundant antagonistic microorganisms in disease-suppressive soils against tobacco bacterial wilt. [ABSTRACT FROM AUTHOR]

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Potato is considered a staple food in the world and its production is limited by the presence of bacterial wilt (bw) disease caused by Ralstonia solanacearum. Host resistance is the most sustainable and cost-effective strategy to manage bw, although resistant commercial potato cultivars are not yet available. Our group incorporated the efr receptor of Arabidopsis thaliana (atefr), which recognizes the elongation factor Tu, preserved in bacteria, triggering an immune response. atefr was tested in two genetic backgrounds: a susceptible commercial cultivar (inia Iporá) and a breeding clone with partial resistance introgressed from Solanum commersonii. In this work, the effect of the atefr receptor on bw resistance was evaluated for the first time, using conditions resembling natural field infection. In addition, the colonization patterns of wild-type and transgenic lines were compared by using luminescent and fluorescent R. solanacearum reporter strains. Both approaches showed a delay and a decrease in the severity of wilting symptoms in the atefr-transformed genotypes. Differential colonization patterns were observed, revealing a higher bacterial development in the non-transformed plants. This atefr effect seems more pronounced in the interspecific breeding line, possibly leading to a more effective activation of the plant immune system.

Bacterial wilt is caused by Ralstonia solanacearum is a most destructive and prevalent disease in worldwide causing enormous yield losses in tomato. In the present study, eight parental lines along with their twenty-eight F1 hybrids were evaluated against bacterial wilt. Among the parents, two parents CHFT-71 and CHFT-50, showed moderately resistant against wilt. However, none of the parents recorded a highly resistant and immune to the bacterial wilt. Only single cross, CHFT-71×CHFT-50 showed a wilt resistant and four hybrids namely, CHFT-77×CHFT-71, CHFT-79×CHFT-50, CHFT-60×CHFT-50 and H-86×CHFT-50 were found to be moderately resistant against wilt. These five hybrids can be recommended for further evaluation. The promising parental lines can be utilised for the improvement of varieties and development of wilt resistant F1’s through hybrid breeding programmes.

Bacterial wilt caused by Ralstonia solanacearum is an important disease of tomatoes. The grafting method with a resistant rootstock variety is a reliable control technique and has been carried out in many countries. Healthy tomato seedling are needed for grafting so this study was aimed to find a good seedling medium to grow the upper stem and rootstock of tomato for grafting and to know the effect of seed media on the intensity of bacterial wilt). Tomato seeds were grown in coir and rice husk charcoal with ratio A (10:0), B (9:1), C (8:2), D (7:3), E (6:4), F (5:5) (coir : rice husk charcoal, v/v). Seedlings were watered regularly and after seedlings aged 21 days, the grafting was conducted. Agronomic observations were justified by measuring germination style, plant height, leaf area, fresh mass and dry mass of plants at 21 days after planting. Tomato seedlings were grafted by using H-7996 variety as a rootstock and Servo variety as an upper stem. Tomato seedlings were transplanted into polybags and then inoculated with R. solanacearum (108 CFU/ml). The disease intensity and AUDPC (Area Under Disease Progress Curve) were recorded. The results revealed that the media with a ratio of 8:2 (coir : rice husk charcoal, v/v) is the best medium for growing tomatoes and the grafting treatment could reduce the disease intensity of bacterial wilt by 40% compared with non grafted treatment and reduce 16% disease intensity compared to the grafted treatment with seedling sown in other ratio media at the last observation.

Ralstonia solanacearum, the causal agent of bacterial wilt and brown rot disease, is one of the major pathogens of solanaceous crops, including potato, around the globe. Biovar 2T (phylotype II/sequevar 25) of R. solanacearum is adapted to tropical lowlands and is only reported in South America and Iran. Thus far, no genome resource of the biovar 2T of the pathogen has been available. Here, we present the near-complete genome sequences of the biovar 2T strain CFBP 8697 as well as strain CFBP 8695 belonging to biovar 2 race 3, both isolated from potato in Iran. The genomic data of biovar 2T will extend our understanding of the virulence features of R. solanacearum and pave the way for research on biovar 2T functional and interaction genetics.

Abstract Background There is an urgent need to discover biocontrol agents to control bacterial wilt. This study reports on a new lipopeptide-producing biocontrol strain FJAT-46737 and explores its lipopeptidic compounds, and this study investigates the antagonistic effects of these compounds. Results Based on a whole genome sequence analysis, the new strain FJAT-46737 was identified as Bacillus velezensis, and seven gene clusters responsible for the synthesis of bioactive secondary metabolites in FJAT-46737 were predicted. The antimicrobial results demonstrated that FJAT-46737 exhibited broad-spectrum antimicrobial activities in vitro against three bacteria and three fungi. Pot experiments showed that the control efficiencies for tomato bacterial wilt of the whole cultures, the 2-fold diluted supernatants and the crude lipopeptide of FJAT-46737 were 66.2%, 82.0%, and 96.2%, respectively. The above results suggested that one of the antagonistic mechanisms of FJAT-46737 was the secretion of lipopeptides consisting of iturins, fengycins and surfactins. The crude lipopeptides had significant antagonistic activities against several pathogens (including Ralstonia solanacearum, Escherichia coli and Fusarium oxysporum) and fengycins were the major antibacterial components of the lipopeptides against R. solanacearum in vitro. Furthermore, the rich organic nitrogen sources (especially yeast extracts) in the media promoted the production of fengycin and surfactin by FJAT-46737. The secretion of these two lipopeptides was related to temperature fluctuations, with the fengycin content decreasing by 96.6% and the surfactins content increasing by 59.9% from 20 °C to 40 °C. The optimal temperature for lipopeptide production by FJAT-46737 varied between 20 °C and 25 °C. Conclusions The B. velezensis strain FJAT-46737 and its secreted lipopeptides could be used as new sources of potential biocontrol agents against several plant pathogens, and especially the bacterial wilt pathogen R. solanacearum.

Ralstonia solanacearum (Rso) is a causal agent of bacterial wilt in Solanaceae crops worldwide including Republic of Korea. Rso virulence predominantly relies on type III secreted effectors (T3Es). However, only a handful of Rso T3Es have been characterized. In this study, we investigated subcellular localization of and manipulation of plant immunity by 8 Rso T3Es predicted to harbor a nuclear localization signal (NLS). While 2 of these T3Es elicited cell death in both Nicotiana benthamiana and N. tabacum, only one was dependent on suppressor of G2 allele of skp1 (SGT1), a molecular chaperone of nucleotide-binding and leucine-rich repeat immune receptors. We also identified T3Es that differentially regulate flg22-induced reactive oxygen species production and gene expression. Interestingly, several of the NLS-containing T3Es translationally fused with yellow fluorescent protein accumulated in subcellular compartments other than the cell nucleus. Our findings bring new clues to decipher Rso T3E function in planta.

Silicon (Si) can increase plant resistance against bacterial wilt caused by Ralstonia solanacearum and enhance plant immune response. However, whether Si alleviates soil-borne disease stress through altering soil microbial community component and diversity is not clear. In this study, effects of Si application under R. solanacearum inoculation with or without plant on soil bacterial and fungal communities were investigated through high-throughput pyrosequencing technique. The results showed that Si addition significantly reduced bacterial wilt incidence. However, Si did not reduce the amount of R. solanacearum in rhizosphere soil. Principal components analysis showed that soil microbial community composition was strongly influenced by Si addition. Total 63.7% bacterial operational taxonomic units (OTUs) and 43.8% fungal OTUs were regulated by Si addition regardless of the presence of tomato plants, indicating the independent effects of Si on soil microbial community. Si-added soil harbored a lower abundance of Fusarium, Pseudomonas, and Faecalibacterium. Our finding further demonstrated that exogenous Si could significantly influence soil microbial community component, and this may provide additional insight into the mechanism of Si-enhanced plant resistance against soil-borne pathogens.

Pogostemon cablin (patchouli), an important medicinal and aromatic plant, is widely used in traditional Chinese medicine as well as in perfume industry. Patchouli plants are susceptible to bacterial wilt disease, which causes significant economic losses by reduction in yield and quality of the plant products. However, few studies focus on the pathogens causing bacterial wilt on patchouli. In this study, strain Pa82 was isolated from diseased patchouli plants with typical bacterial wilt symptoms in Guangdong province, China, and was confirmed to be a highly virulent pathogen of patchouli bacterial wilt. Comparative sequence analysis of 16S rRNA gene showed that the strain was closely related to Kosakonia sp. CCTCC M2018092 (99.9% similarity) and Kosakonia cowanii Esp_Z (99.8% similarity). Moreover, phylogenetic tree based on 16S rRNA gene sequences showed that the strain was affiliated with genus Kosakonia. Further, the whole genome of strain Pa82 was sequenced, and the sequences were assembled and annotated. The complete genome of the strain consists of one chromosome and three plasmids. Average nucleotide identity (ANI) and phylogenetic analysis revealed that the strain belongs to Kosakonia cowanii (designated Kosakonia cowanii Pa82). Virulence-related genes of the strain involved in adherence, biofilm formation, endotoxin and other virulence factors were predicted. Among them, vgrG gene that encodes one of the type VI secretion system components was functionally validated as a virulence factor in Kosakonia cowanii Pa82 through construction of Tn5 insertion mutants and identification of mutant defective in virulence.

Ralstonia solanacearum is the causal agent of bacterial wilt, one of the most destructive diseases of solanaceous plants, affecting staple crops worldwide. The bacterium survives in water, soil, and other reservoirs, and is difficult to control. In this sense, the use of three specific lytic R. solanacearum bacteriophages was recently patented for bacterial wilt biocontrol in environmental water and in plants. To optimize their applications, the phages and the bacterium need to be accurately monitored and quantified, which is laborious and time-consuming with biological methods. In this work, primers and TaqMan probes were designed, and duplex and multiplex real-time quantitative PCR (qPCR) protocols were developed and optimized for the simultaneous quantification of R. solanacearum and their phages. The quantification range was established from 108 to 10 PFU/mL for the phages and from 108 to 102 CFU/mL for R. solanacearum. Additionally, the multiplex qPCR protocol was validated for the detection and quantification of the phages with a limit ranging from 102 targets/mL in water and plant extracts to 103 targets/g in soil, and the target bacterium with a limit ranging from 103 targets/mL in water and plant extracts to 104 targets/g in soil, using direct methods of sample preparation.

The foundation of effectively predicting plant disease in the early stage using deep learning algorithms is ideal for addressing food insecurity, inevitably drawing researchers and agricultural specialists to contribute to its effectiveness. The input preprocessor, abnormalities of the data (i.e., incomplete and nonexistent features, class imbalance), classifier, and decision explanation are typical components of a plant disease detection pipeline based on deep learning that accepts an image as input and outputs a diagnosis. Data sets related to plant diseases frequently display a magnitude imbalance due to the scarcity of disease outbreaks in real field conditions. This study examines the effects of several preprocessing methods and class imbalance approaches and deep learning classifiers steps in the pipeline for detecting plant diseases on our data set. We notably want to evaluate if additional preprocessing and effective handling of data inconsistencies in the plant disease pipeline may considerably assist deep learning classifiers. The evaluation’s findings indicate that contrast limited adaptive histogram equalization (CLAHE) combined with image sharpening and generative adversarial networks (GANs)-based approach for resampling performed the best among the preprocessing and resampling techniques, with an average classification accuracy of 97.69% and an average F1-score of 97.62% when fed through a ResNet-50 as the deep learning classifier. Lastly, this study provides a general workflow of a disease detection system that allows each component to be individually focused on depending on necessity.

Casuarina equisetifolia is crucial in protecting coastal regions of China against typhoon attacks but has faced a substantial challenge due to wilt disease caused by pathogens of the Ralstonia solanacearum species complex (RSSC). Although the initial outbreak of Casuarina wilt in the 1970s was effectively controlled by disease-resistant C. equisetifolia varieties, the disease has recently re-emerged in coastal regions of Guangdong. In this study, we report the isolation, characterization, and comparative genomic analysis of 11 RSSC strains from diseased C. equisetifolia at various locations along the coast of Guangdong. Phylogenomic analysis showed that the strains were closely related and clustered with phylotype I strains previously isolated from peanuts. Single-gene-based analysis further suggested these strains could be derived from strains present in Guangdong since the 1980s, indicating a historical context to their current pathogenicity. Casuarina -isolated strains exhibited notably higher virulence against C. equisetifolia and peanuts than the representative RSSC strains GMI1000 and EP1, suggesting host-specific adaptations that possibly contributed to the recent outbreak. Comparative genomic analysis among RSSC strains revealed a largely conserved genome structure and high levels of conservation in gene clusters encoding extracellular polysaccharide biosynthesis, secretion systems, and quorum sensing regulatory systems. However, we also found a number of unique genes in the Casuarina -isolated strains that were absent in GMI1000 and EP1, and vice versa, pointing to potential genetic factors underpinning their differential virulence. These unique genes offer promising targets for future functional studies. Overall, our findings provide crucial insights into the RSSC pathogens causing Casuarina wilt in Guangdong, guiding future efforts in disease control and prevention., Competing Interests: The author(s) declare no conflict of interest.

PAMP-triggered immunity (PTI) is the first layer of plant defense response that occurs on the plant plasma membrane. Recently, the application of a rhizobacterium, Bacillus amyloliquefaciens strain PMB05, has been demonstrated to enhance flg22 Pst - or harpin-triggered PTI response such as callose deposition. This PTI intensification by PMB05 further contributes to plant disease resistance to different bacterial diseases. Under the demand for rapid and large-scale screening, it has become critical to establish a non-staining technology to identify microbial strains that can enhance PTI responses. Firstly, we confirmed that the expression of the GSL5 gene, which is required for callose synthesis, can be enhanced by PMB05 during PTI activation triggered by flg22 or PopW (a harpin from Ralstonia solanacearum ). The promoter region of the GSL5 gene was further cloned and fused to the coding sequence of gfp . The constructed fragments were used to generate transgenic Arabidopsis plants through a plant transformation vector. The transgenic lines of At GSL5 -GFP were obtained. The analysis was performed by infiltrating flg22 Pst or PopW in one homozygous line, and the results exhibited that the green fluorescent signals were observed until after 8 h. In addition, the PopW-induced fluorescent signal was significantly enhanced in the co-treatment with PMB05 at 4 h after inoculation. Furthermore, by using At GSL5 -GFP to analyze 13 Bacillus spp. strains, the regulation of PopW-induced fluorescent signal was observed. And, the regulation of these fluorescent signals was similar to that performed by callose staining. More importantly, the Bacillus strains that enhance PopW-induced fluorescent signals would be more effective in reducing the occurrence of bacterial wilt. Taken together, the technique by using At GSL5 -GFP would be a promising platform to screen plant immunity-intensifying microbes to control bacterial wilt.

In Malaysia, bananas ( Musa spp.) are the second most cultivated fruit and the fourth most cultivated fruit in terms of export revenue. In October 2018, about 5.0 out of 6.6 hectares of a banana plantation located in Teluk Intan, Malaysia, was impacted by an outbreak of banana disease. The onset of bacterial wilt symptoms is characterized by initial leaf wilting, followed by the subsequent withering of the entire plant during later stages, fruit stalk and fruit pulp discoloration, fruit rotting, and pseudostem necrosis. The diseased banana's symptomatic pseudostems and fruit pulps were surface-sterilised in 70% ethanol for 30 s, followed by 2% NaClO for 3 min, rinsed three times in sterilised water, and cut into small pieces approximately 5 mm 2 in size. The tissues were macerated in a sterilised 0.85% NaCl solution for 5 min, and the resulting suspension was streaked onto nutrient agar, followed by incubation at 28°C for 2 days. After incubation, bacterial colonies with five unique morphological characteristics were observed. Two colonies of each unique morphological type were randomly chosen and subjected to preliminary bacterial identification by 16S rRNA gene sequencing. Based on BLASTn analysis, the five unique morphological types of bacteria were preliminarily identified as Enterobacter cloacae , Citrobacter farmeri , Klebsiella variicola , Kosakonia radicincitans , and Phytobacter ursingii . Previous reports identified K. variicola and K. radicincitans as banana pathogens, but Malaysia has yet to report the former. The amplified partial 16S rDNA sequences of both K. variicola isolates (designated as UTAR-BC1 and UTAR-BC2; GenBank accession numbers: PP531448 and PP531460, respectively), which were chosen to be the focus of this study, exhibited complete similarity to each other and were 100% identical (1426/1426 identity and 1420/1420 identity, respectively) to K. variicola (CP026013.1). To verify the identity of the bacterial isolate, three housekeeping genes, namely, inf B (PP538994), rpo B (PP538995), and gyr B (PP538996) of UTAR-BC1, were amplified, sequenced, and subjected to multilocus phylogenetic analysis via the neighbour-joining method (1,000 bootstrap values). Phylogenetic analysis revealed that UTAR-BC1 belongs to the K. variicola clade. A pathogenicity assay of UTAR-BC1 was conducted on 4-month-old healthy banana plantlets (cv. Nangka) using the pseudostem injection method (Tripathi et al., 2008). First, UTAR-BC1 was grown overnight in nutrient broth and then adjusted to 10 8 CFU/ml in a sterile 10 mM MgCl 2 solution. A total volume of 100 µL of the bacterial suspension was injected into the pseudostem of five healthy banana plantlets via a syringe with a needle. Control plants were mock-inoculated with a sterile 10 mM MgCl 2 solution. The experiments were replicated thrice and inoculated plants were maintained at room temperature with natural sunlight and humidity, which resembled the field conditions. Two months after inoculation, all of the UTAR-BC1 inoculated spots of banana plantlets showed severe necrosis, while the banana leaves showed symptoms of wilted appearance, whereas the control plants remained symptomless. The reisolated pathogen from 90% of the symptomatic pseudostems and leaf blades shares the same morphological and molecular features as UTAR-BC1, thus fulfilling Koch's postulates. Previously, K. variicola has been reported to be a banana pathogen causing rhizome rot in India (Loganathan et al., 2021), plantain soft rot in Haiti (Fulton et al. 2020), and sheath rot and bulb rot in China (Sun et al., 2023; Jiang et al., 2024). To the best of our knowledge, this is the first report of bacterial wilt disease in bananas attributed to K. variicola in Malaysia. This finding will facilitate the surveillance of K. variicola as an emerging pathogen in banana plants in this region, thereby safeguarding the country's food security and promoting socio-economic growth.

Tobacco bacterial wilt is a highly destructive soilborne disease caused by the Ralstonia solanacearum species complex, exhibiting a significant risk to global flue-cured tobacco cultivation and resulting in substantial economic loss. In this study, 77 isolates were collected from three prominent flue-cured tobacco cultivation areas in Fujian, China (Nanping, Sanming, and Longyan), in 2021 and 2022. The isolated strains were classified through phylotype-specific multiplex polymerase chain reaction (Pmx-PCR) and physiological tests. The analysis showed that all the strains were associated with phylotype I, race 1, and biovar III. Subsequent phylogenetic analysis using partial egl gene sequences classified the 77 isolates into 5 distinct sequevars: 13, 15, 16, 17, and 34. Notably, a remarkable predominance of sequevar 15 was observed in Fujian Province, while sequevar 16 was first reported on tobacco in China, which was identified in other plants, expanding the understanding of its host range and distribution in the country. In addition, a Streptomyces strain extracted from the rhizosphere soil of tobacco was found to inhibit the growth of multiple sequevars of tobacco R. solanacearum , indicating its broad-spectrum antagonistic properties. Furthermore, pot experiments showed that the strain St35 effectively controlled tobacco bacterial wilt. The isolate St35 was conclusively identified as Streptomyces gancidicus according to the morphological and genetic features. In summary, the present study demonstrated the genetic diversity and distribution of tobacco R. solanacearum strains in the Fujian province of China, as well as the identification of a candidate biological control agent for the management of tobacco bacterial wilt., Competing Interests: The author(s) declare no conflict of interest.