Endophytic bacterial communities in tomato plants with differential resistance to Ralstonia solanacearum

ABSTRACT Ralstonia solanacearum, a widely distributed and economically important plant pathogen, invades the roots of diverse plant hosts from the soil and aggressively colonizes the xylem vessels, causing a lethal wilting known as bacterial wilt disease. By examining bacteria from the xylem vessels of infected plants, we found thatR. solanacearum is essentially nonmotile in planta, although it can be highly motile in culture. To determine the role of pathogen motility in this disease, we cloned, characterized, and mutated two genes in the R. solanacearum flagellar biosynthetic pathway. The genes for flagellin, the subunit of the flagellar filament (fliC), and for the flagellar motor switch protein (fliM) were isolated based on their resemblance to these proteins in other bacteria. As is typical for flagellins, the predicted FliC protein had well-conserved N- and C-terminal regions, separated by a divergent central domain. The predicted R. solanacearum FliM closely resembled motor switch proteins from other proteobacteria. Chromosomal mutants lackingfliC or fliM were created by replacing the genes with marked interrupted constructs. Since fliM is embedded in the fliLMNOPQR operon, the aphAcassette was used to make a nonpolar fliM mutation. Both mutants were completely nonmotile on soft agar plates, in minimal broth, and in tomato plants. The fliC mutant lacked flagella altogether; moreover, sheared-cell protein preparations from the fliC mutant lacked a 30-kDa band corresponding to flagellin. The fliM mutant was usually aflagellate, but about 10% of cells had abnormal truncated flagella. In a biologically representative soil-soak inoculation virulence assay, both nonmotile mutants were significantly reduced in the ability to cause disease on tomato plants. However, the fliC mutant had wild-type virulence when it was inoculated directly onto cut tomato petioles, an inoculation method that did not require bacteria to enter the intact host from the soil. These results suggest that swimming motility makes its most important contribution to bacterial wilt virulence in the early stages of host plant invasion and colonization.

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NtRNF217, Encoding a Putative RBR E3 Ligase Protein of Nicotiana tabacum, Plays an Important Role in the Regulation of Resistance to Ralstonia solanacearum Infection

International Journal of Molecular Sciences ◽

10.3390/ijms22115507 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5507

Author(s):

Ying Liu ◽

Yuanman Tang ◽

Xi Tan ◽

Wei Ding

Keyword(s):

Nicotiana Tabacum ◽

Ralstonia Solanacearum ◽

Plant Immunity ◽

E3 Ligase ◽

Marker Genes ◽

Susceptible Cultivar ◽

Wild Type ◽

Disease Symptoms ◽

Short Time ◽

Resistance To Ralstonia Solanacearum

E3 ubiquitin ligases, the most important part of the ubiquitination process, participate in various processes of plant immune response. RBR E3 ligase is one of the E3 family members, but its functions in plant immunity are still little known. NtRNF217 is a RBR E3 ligase in tobacco based on the sequence analysis. To assess roles of NtRNF217 in tobacco responding to Ralstonia solanacearum, overexpression experiments in Nicotiana tabacum (Yunyan 87, a susceptible cultivar) were performed. The results illuminated that NtRNF217-overexpressed tobacco significantly reduced multiplication of R. solanacearum and inhibited the development of disease symptoms compared with wild-type plants. The accumulation of H2O2 and O2− in NtRNF217-OE plants was significantly higher than that in WT-Yunyan87 plants after pathogen inoculation. The activities of CAT and SOD also increased rapidly in a short time after R. solanacearum inoculation in NtRNF217-OE plants. What is more, overexpression of NtRNF217 enhanced the transcript levels of defense-related marker genes, such as NtEFE26, NtACC Oxidase, NtHIN1, NtHSR201, and NtSOD1 in NtRNF217-OE plants after R. solanacearum inoculation. The results suggested that NtRNF217 played an important role in regulating the expression of defense-related genes and the antioxidant enzymes, which resulted in resistance to R. solanacearum infection.

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Interactions with Hosts at Cool Temperatures, Not Cold Tolerance, Explain the Unique Epidemiology of Ralstonia solanacearum Race 3 Biovar 2

Phytopathology ◽

10.1094/phyto-99-10-1127 ◽

2009 ◽

Vol 99 (10) ◽

pp. 1127-1134 ◽

Cited By ~ 43

Author(s):

Annett Milling ◽

Fanhong Meng ◽

Timothy P. Denny ◽

Caitilyn Allen

Keyword(s):

North American ◽

Ralstonia Solanacearum ◽

Potato Tubers ◽

In Planta ◽

Plant Host ◽

Tomato Plants ◽

North American Strain ◽

Tropical Highlands ◽

American Strain ◽

Warm Temperate

Most strains of the bacterial wilt pathogen Ralstonia solanacearum are tropical, but race 3 biovar 2 (R3bv2) strains can attack plants in temperate zones and tropical highlands. The basis of this distinctive ecological trait is not understood. We compared the survival of tropical, R3bv2, and warm-temperate North American strains of R. solanacearum under different conditions. In water at 4°C, North American strains remained culturable the longest (up to 90 days), whereas tropical strains remained culturable for the shortest time (≈40 days). However, live/dead staining indicated that cells of representative strains remained viable for >160 days. In contrast, inside potato tubers, R3bv2 strain UW551 survived >4 months at 4°C, whereas North American strain K60 and tropical strain GMI1000 were undetectable after <70 days in tubers. GMI1000 and UW551 grew similarly in minimal medium at 20 and 28°C and, although both strains wilted tomato plants rapidly at 28°C, UW551 was much more virulent at 20°C, killing all inoculated plants under conditions where GMI100 killed just over half. Thus, differences among the strains in the absence of a plant host were not predictive of their behavior in planta at cooler temperatures. These data indicate that interaction with plants is required for expression of the temperate epidemiological trait of R3bv2.

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Selection and Inheritance of Tomato Resistance against Ralstonia solanacearum

Jurnal Perlindungan Tanaman Indonesia ◽

10.22146/jpti.35464 ◽

2019 ◽

Vol 23 (1) ◽

pp. 61

Author(s):

Isna Maulida ◽

Rudi Hari Murti ◽

Triwidodo Arwiyanto

Keyword(s):

Bacterial Wilt ◽

Ralstonia Solanacearum ◽

Dominant Gene ◽

Tomato Plants ◽

Breeding Lines ◽

Back Cross ◽

Inheritance Patterns ◽

Randomized Design ◽

Resistant Varieties ◽

Resistant Lines

Ralstonia solanacearum is a plant pathogen causes wilting which is a major obstacle in the cultivation of tomato plants. In plant breeding, knowledge of the source of resistance genes and inheritance patterns is important in the development of bacterial wilt resistant varieties. This study aimed to obtain bacterial wilt resistant lines and to find out the inheritance pattern of tomato resistance to bacterial wilt. Selection of resistant plant involved the selected breeding lines from irradiation and crossing collections of the Genetic Laboratory, Faculty of Agriculture, Universitas Gadjah Mada. Introduced lines of H-7996 and F1 Permata and Timoti were used as a control. H-7996 as resistant parents and GM2 as susceptible parents, and their offspring include F1 GM2 x H-7996, F1 reciprocal, F2, Back Cross 1 (F1 x GM2), and Back Cross 2 (F1 x H-7996) used in testing inheritance patterns. Inoculation was carried out 1 week after planting by pouring 100 ml of water suspension of R. solanacarum (108 cfu/ml) on the roots. Completely Randomized Design (CRD) was used in this experiment. The scoring observation was carried out every week for one month. This study showed that Permata as a control was the most resistant, while Timoti and H-7996 were medium resistant. The CLN, G6, G8, and G7 lines were susceptible medium, yet only G8 and G7 with the smallest percentage of disease intensity and not significantly different than Timoti. The resistance gene to bacterial wilt on H-7996 was controlled by genes in the cell nucleus with additive-dominant gene action. Resistance to bacteria has a moderate level of heritability.

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Resistance and Susceptibility of Arabidopsis thaliana to Bacterial Wilt Caused by Ralstonia solanacearum

Phytopathology ◽

10.1094/phyto.1998.88.4.330 ◽

1998 ◽

Vol 88 (4) ◽

pp. 330-334 ◽

Cited By ~ 15

Author(s):

Chang-Hsien Yang ◽

Gan-Der Ho

Keyword(s):

Arabidopsis Thaliana ◽

Bacterial Wilt ◽

Ralstonia Solanacearum ◽

Bacterial Colonization ◽

Plant Diseases ◽

Infection Site ◽

Bacterial Isolation ◽

Tomato Plants ◽

Different Strains ◽

Growth Of Bacteria

Tomato bacterial wilt caused by Ralstonia solanacearum is a model system for studying plant-bacterial interactions, because it is genetically one of the best characterized plant diseases. We demonstrate here that four different strains of R. solanacearum, two from radishes (Rd4 and Rd15) and two from tomato (Ps21 and Ps95), can infect 27 different ecotypes of Arabidopsis thaliana, causing different responses. All ecotypes tested were highly susceptible to strain Rd15, which caused symptoms similar to those observed in tomato plants. For example, leaf drooping and discoloration developed just 3 days after inoculation, and plants completely wilted within 1 week. Strains Rd4 and Ps95 were less infectious than Rd15. With these two strains, a variety of disease responses were observed among different ecotypes at 2 weeks after inoculation; both susceptible and resistant ecotypes of A. thaliana were identified. Ps21 was the least infectious of the four strains and caused almost no symptoms in any of the ecotypes of Arabidopsis tested. Direct bacterial isolation and plant skeleton hybridization analysis from infected plants indicated that bacterial colonization was correlated with the severity of symptoms. Growth of bacteria was limited to the infection site in resistant plants, whereas the bacteria spread throughout susceptible plants by 1 week after inoculation.

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Ethylene emission and PR protein synthesis in ACC deaminase producing Methylobacterium spp. inoculated tomato plants (Lycopersicon esculentum Mill.) challenged with Ralstonia solanacearum under greenhouse conditions

Plant Physiology and Biochemistry ◽

10.1016/j.plaphy.2013.03.002 ◽

2013 ◽

Vol 67 ◽

pp. 95-104 ◽

Cited By ~ 34

Author(s):

Woojong Yim ◽

Sundaram Seshadri ◽

Kiyoon Kim ◽

Gillseung Lee ◽

Tongmin Sa

Keyword(s):

Protein Synthesis ◽

Lycopersicon Esculentum ◽

Ralstonia Solanacearum ◽

Acc Deaminase ◽

Tomato Plants ◽

Pr Protein ◽

Lycopersicon Esculentum Mill ◽

Ethylene Emission

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Influence of Pythium oligandrum on the bacterial communities that colonize the nutrient solutions and the rhizosphere of tomato plants

Canadian Journal of Microbiology ◽

10.1139/w2012-092 ◽

2012 ◽

Vol 58 (9) ◽

pp. 1124-1134 ◽

Cited By ~ 10

Author(s):

J. Vallance ◽

F. Déniel ◽

G. Barbier ◽

L. Guerin-Dubrana ◽

N. Benhamou ◽

...

Keyword(s):

Genetic Structure ◽

Root System ◽

Quantitative Pcr ◽

Bacterial Communities ◽

Single Strand Conformation Polymorphism ◽

Pythium Oligandrum ◽

Tomato Plants ◽

Cropping Season ◽

Nutrient Solutions ◽

Conformation Polymorphism

The influence exerted by the biocontrol oomycete Pythium oligandrum on the bacterial populations proliferating in the rhizosphere of tomato plants grown in a hydroponic system and in the circulating solutions is studied in the present experiment. Quantitative PCR and single-strand conformation polymorphism were used to investigate the genetic structure and dynamics of the bacterial communities colonizing the root systems and the various circulating solutions. Quantitative PCR assays showed that bacteria heavily colonized the rhizosphere of tomato plants with, however, no significant density changes throughout the cultural season (April–September). Single strand conformation polymorphism fingerprints revealed the occurrence of transient perturbations in the rhizospheric indigenous bacterial communities following P. oligandrum introduction in the root system of plants. This effect was, however, transient and did not persist until the end of the cropping season. Interestingly, the genetic structure of the bacterial microflora colonizing either the roots or the nutrient solutions evolved throughout the cropping season. This temporal evolution occurred whatever the presence and persistence of P. oligandrum in the rhizosphere. Evidence is also provided that bacterial microflora that colonize the root system are different from the ones colonizing the circulating solutions. The relationships between these 2 microflora (at the root and solution levels) are discussed.

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Biological control of bacterial wilt of tomato caused by Ralstonia solanacearum using Pseudomonas species isolated from the rhizosphere of tomato plants

Archives of Phytopathology and Plant Protection ◽

10.1080/03235408.2020.1715756 ◽

2020 ◽

Vol 53 (1-2) ◽

pp. 1-16 ◽

Cited By ~ 4

Author(s):

Ayomide F. Mohammed ◽

Adejare R. Oloyede ◽

Adebola O. Odeseye

Keyword(s):

Biological Control ◽

Bacterial Wilt ◽

Ralstonia Solanacearum ◽

Tomato Plants ◽

Pseudomonas Species

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Pyramiding Unmarked Deletions in Ralstonia solanacearum Shows That Secreted Proteins in Addition to Plant Cell-Wall-Degrading Enzymes Contribute to Virulence

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-18-1296 ◽

2005 ◽

Vol 18 (12) ◽

pp. 1296-1305 ◽

Cited By ~ 77

Author(s):

Huanli Liu ◽

Shuping Zhang ◽

Mark A. Schell ◽

Timothy P. Denny

Keyword(s):

Cell Wall ◽

Plant Cell ◽

Ralstonia Solanacearum ◽

Plant Cell Wall ◽

Secreted Proteins ◽

Cellulolytic Enzymes ◽

Wild Type ◽

Cell Wall Degrading Enzymes ◽

Tomato Plants ◽

Degrading Enzymes

Ralstonia solanacearum, like many phytopathogenic bacteria, makes multiple extracellular plant cell-wall-degrading enzymes (CWDE), some of which contribute to its ability to cause wilt disease. CWDE and many other proteins are secreted to the milieu via the highly conserved type II protein secretion system (T2SS). R. solanacearum with a defective T2SS is weakly virulent, but it is not known whether this is due to absence of all the CWDE or the loss of other secreted proteins that contribute to disease. These alternatives were investigated by creating mutants of wild-type strain GMI1000 lacking either the T2SS or up to six CWDE and comparing them for virulence on tomato plants. To create unmarked deletions, genomic regions flanking the target gene were polymerase chain reaction (PCR)-amplified, were fused using splice overlap extension PCR, were cloned into a suicide plasmid harboring the sacB counter-selectable marker, and then, were site-specifically introduced into the genome. Various combinations of five deletions (δpehA, δpehB, δpehC, δpme, and δegl) and one inactivated allele (cbhA::aphA-3) resulted in 15 mutants missing one to six CWDE. In soil-drench inoculation assays, virulence of mutants lacking only pectic enzymes (PehA, PehB, PehC, and Pme) was not statistically different from GMI1000, but all the mutants lacking one or both cellulolytic enzymes (Egl or CbhA) wilted plants significantly more slowly than did the wild type. The GMI-6 mutant that lacks all six CWDE was more virulent than the mutant lacking only its two cellulolytic enzymes, and both were significantly more virulent than the T2SS mutant (GMI-D). Very similar results were observed in wounded-petiole inoculation assays, so GMI-6 and GMI-D appear to be less capable of colonizing tomato tissues after invasion. Because the T2SS mutant was much less virulent than the sixfold CWDE mutant, we conclude that other secreted proteins contribute substantially to the ability of R. solanacearum GMI1000 to systemically colonize tomato plants.

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