Effects of the pseudomonad phytotoxin coronatine on tomato leaf structure and ultrastructure

1995 ◽  
Vol 53 ◽  
pp. 862-863
Author(s):  
D.A. Palmer ◽  
C.L. Bender
Keyword(s):  
Pseudomonas Syringae ◽  
Membrane Integrity ◽  
Leaf Tissue ◽  
Cell Number ◽  
Cell Ultrastructure ◽  
Response To Treatment ◽  
Prominent Symptom ◽  
Chlorophylls A And B ◽  
Cell Dimensions ◽  
Structure And Ultrastructure

Coronatine is a non-host-specific phytotoxin produced by several members of the Pseudomonas syringae group of pathovars. The toxin acts as a virulence factor in P. syringae pv. tomato, allowing the organism to multiply to a higher population density and develop larger lesions than mutant strains unable to produce the toxin. The most prominent symptom observed in leaf tissue treated with coronatine is an intense spreading chlorosis; this has been attributed to a loss of chlorophylls a and b in tobacco. Coronatine's effects on membrane integrity and cell ultrastructure have not been previously investigated. The present study describes changes in tomato leaves in response to treatment with purified coronatine, infection by a coronatine-producing strain of P. syringae pv. tomato, and infection by a cor" mutant.In contrast to H2O-treated tissue, coronatine-treated tissue showed a diffuse chlorosis extending approximately 5 mm from the inoculation site. Leaf thickness, cell number, and cell dimensions were similar for both healthy and coronatine-treated, chlorotic tissue; however, the epidermal cell walls were consistently thicker in coronatine-treated leaves (Figs, la and lb).

An In Vitro Cytotoxicity Assay Using Rat Hepatocytes and MTT and Coomassie Blue Dye as Indicators

1991 ◽  
Vol 19 (3) ◽  
pp. 352-360
Author(s):  
Kazuhiko Otoguro ◽  
Kanki Komiyama ◽  
Satoshi Ωmura ◽  
Charles A. Tyson
Keyword(s):  
Mtt Assay ◽  
Membrane Integrity ◽  
Cell Number ◽  
Cellular Protein ◽  
Blue Dye ◽  
Tetrazolium Salt ◽  
Sprague Dawley ◽  
Lactate Dehydrogenase Activity ◽  
Coomassie Blue

Isolated hepatocytes from male Sprague-Dawley rats suspended in culture medium supplemented with either 0.2 or 2% bovine serum albumin (BSA) were allowed to attach to collagen coated 96-well dishes. Ten test chemicals from the MEIC list and salicylic acid were added individually to the dishes, and at the end of 24 and 48 hours, cytotoxicity was determined by measuring MTT (tetrazolium salt) reduction (mitochondrial integrity) and total cellular protein using Coomassie blue dye (reflecting cell number). Total cellular lactate dehydrogenase activity was also determined in some experiments, as an indicator of plasma membrane integrity. The relative toxicities of the test chemicals were quantified by the estimation of EC10, EC20 and EC50 values for each parameter. Except for one chemical, digoxin, in the MTT assay, cytotoxic potency increased with incubation time. The hepatocytes tended to be more sensitive to the chemicals in medium containing 0.2% BSA than in medium containing 2% BSA. Simple linear regression analyses of the log transformed data from the MTT assay versus log oral LD50 in rats for the test chemicals gave the best results using EC10 at 24 hours (r2 = 0.86). With protein as the cytotoxic indicator, the best results were obtained with EC values in the medium containing 2% BSA, again at 24 hours (r2 = 0.83). These results suggest that the MTT and Coomassie blue dye assays could be useful indicators for testing the cytotoxic potential of chemicals in rat hepatocyte cultures.

scholarly journals Venous and Arterial Endothelial Cells from Human Umbilical Cords: Potential Cell Sources for Cardiovascular Research

2021 ◽  
Vol 22 (2) ◽  
pp. 978
Author(s):  
Skadi Lau ◽  
Manfred Gossen ◽  
Andreas Lendlein ◽  
Friedrich Jung
Keyword(s):  
Endothelial Cells ◽  
Umbilical Cord ◽  
Membrane Integrity ◽  
Cell Types ◽  
Cell Number ◽  
Human Umbilical Cord ◽  
Cardiovascular Research ◽  
Cardiovascular Devices ◽  
Arterial Endothelial Cells

Although cardiovascular devices are mostly implanted in arteries or to replace arteries, in vitro studies on implant endothelialization are commonly performed with human umbilical cord-derived venous endothelial cells (HUVEC). In light of considerable differences, both morphologically and functionally, between arterial and venous endothelial cells, we here compare HUVEC and human umbilical cord-derived arterial endothelial cells (HUAEC) regarding their equivalence as an endothelial cell in vitro model for cardiovascular research. No differences were found in either for the tested parameters. The metabolic activity and lactate dehydrogenase, an indicator for the membrane integrity, slightly decreased over seven days of cultivation upon normalization to the cell number. The amount of secreted nitrite and nitrate, as well as prostacyclin per cell, also decreased slightly over time. Thromboxane B2 was secreted in constant amounts per cell at all time points. The Von Willebrand factor remained mainly intracellularly up to seven days of cultivation. In contrast, collagen and laminin were secreted into the extracellular space with increasing cell density. Based on these results one might argue that both cell types are equally suited for cardiovascular research. However, future studies should investigate further cell functionalities, and whether arterial endothelial cells from implantation-relevant areas, such as coronary arteries in the heart, are superior to umbilical cord-derived endothelial cells.

scholarly journals Expression of an Engineered Cecropin Gene Cassette in Transgenic Tobacco Plants Confers Disease Resistance to Pseudomonas syringae pv. tabaci

1997 ◽  
Vol 87 (5) ◽  
pp. 494-499 ◽  
Author(s):  
Y. Huang ◽  
R. O. Nordeen ◽  
M. Di ◽  
L. D. Owens ◽  
J. H. McBeath
Keyword(s):  
Transgenic Plants ◽  
Pseudomonas Syringae ◽  
Gene Fusion ◽  
Leaf Tissue ◽  
Gene Cassette ◽  
Symptom Development ◽  
Bacterial Inoculum ◽  
Proteinase Inhibitor Ii ◽  
And Control ◽  
Bacterial Concentrations

A chimeric gene fusion cassette, consisting of a secretory sequence from barley α-amylase joined to a modified cecropin (MB39) coding sequence and placed under control of the promoter and terminator from the potato proteinase inhibitor II (PiII) gene, was introduced into tobacco by Agrobacterium-mediated transformation. Transgenic and control plants reacted differently when inoculated with tobacco wildfire pathogen Pseudomonas syringae pv. tabaci at various cell concentrations. With control plants (transformed with a PiII-GUS [β-D-glucuronidase] gene fusion), necrosis was clearly visible in leaf tissue infiltrated with bacterial inoculum levels of 102, 103, 104, 105, and 106 CFU/ml. With MB39-transgenic plants, however, necrosis was observed only in the areas infiltrated with the two highest levels (105 and 106 CFU/ml). No necrosis was evident in areas infiltrated with bacterial concentrations of 104 CFU/ml or less. Bacterial multiplication in leaves of MB39-transgenic plants was suppressed more than 10-fold compared to control plants, and absence of disease symptom development was associated with this growth suppression. We conclude that the pathogen-induced promoter and the secretory sequence were competent elements for transforming a cecropin gene into an effective disease-control gene for plants.

The 73-kb pIAA plasmid increases competitive fitness of Pseudomonas syringae subspecies savastanoi in oleander

1993 ◽  
Vol 39 (7) ◽  
pp. 659-664 ◽  
Author(s):  
Sara E. Silverstone ◽  
David G. Gilchrist ◽  
Richard M. Bostock ◽  
Tsune Kosuge
Keyword(s):  
Population Density ◽  
Pseudomonas Syringae ◽  
Indoleacetic Acid ◽  
Leaf Tissue ◽  
Insertion Mutant ◽  
Wild Type ◽  
In Planta ◽  
Bacterial Strains ◽  
Growth And Survival ◽  
Iaa Production

Pseudomonas syringae subsp. savastanoi causes tumors on olive and oleander by producing the plant growth regulators indoleacetic acid (IAA) and cytokinins following infection of the plant. The contribution of IAA production to the ability of P. syringae subsp. savastanoi to grow and survive in oleander leaf tissue was studied. Bacterial strains differing only with respect to IAA production were characterized. Growth and survival of wild-type and two mutant strains of P. syringae subsp. savastanoi in oleander leaf tissue were monitored by weekly colony counts and IAA plate assays. Growth rate of the three strains in culture and in planta did not differ significantly. However, the wild-type strain reached a higher population density and maintained its maximum density at least 9 weeks longer than either mutant population. An insertion mutant containing the IAA plasmid (pIAA), but incapable of IAA production, did not maintain a higher population density than a strain cured of the IAA plasmid. The pIAA-cured strain maintained a higher population density when coinoculated with an IAA-producing strain than when inoculated alone. These results suggest that IAA production may contribute to the fitness of P. syringae subsp. savastanoi in oleander tissue and that the iaa operon alone may be responsible for the competitive advantage of cells harboring pIAA.Key words: indoleacetic acid, bacterial ecology.

scholarly journals Immunogold Labeling of Hrp Pili of Pseudomonas syringae pv. tomato Assembled in Minimal Medium and In Planta

2001 ◽  
Vol 14 (2) ◽  
pp. 234-241 ◽  
Author(s):  
Wenqi Hu ◽  
Jing Yuan ◽  
Qiao-Ling Jin ◽  
Patrick Hart ◽  
Sheng Yang He
Keyword(s):  
Arabidopsis Thaliana ◽  
Pseudomonas Syringae ◽  
Minimal Medium ◽  
Leaf Tissue ◽  
Hypersensitive Reaction ◽  
Immunogold Labeling ◽  
Structural Protein ◽  
In Planta ◽  
Hrp Genes

Hypersensitive reaction and pathogenicity (hrp) genes are required for Pseudomonas syringae pv. tomato (Pst) DC3000 to cause disease in susceptible tomato and Arabidopsis thaliana plants and to elicit the hypersensitive response in resistant plants. The hrp genes encode a type III protein secretion system known as the Hrp system, which in Pst DC3000 secretes HrpA, HrpZ, HrpW, and AvrPto and assembles a surface appendage, named the Hrp pilus, in hrp-gene-inducing minimal medium. HrpA has been suggested to be the Hrp pilus structural protein on the basis of copurification and mutational analyses. In this study, we show that an antibody against HrpA efficiently labeled Hrp pili, whereas antibodies against HrpW and HrpZ did not. Immunogold labeling of bacteria-infected Arabidopsis thaliana leaf tissue with an Hrp pilus antibody revealed a characteristic lineup of gold particles around bacteria and/or at the bacterium-plant contact site. These results confirm that HrpA is the major structural protein of the Hrp pilus and provide evidence that Hrp pili are assembled in vitro and in planta.

scholarly journals Components of the Pseudomonas syringae Type III Secretion System Can Suppress and May Elicit Plant Innate Immunity

2010 ◽  
Vol 23 (6) ◽  
pp. 727-739 ◽  
Author(s):  
Hye-Sook Oh ◽  
Duck Hwan Park ◽  
Alan Collmer
Keyword(s):  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Leaf Tissue ◽  
Secretion System ◽  
Colony Development ◽  
Plant Responses ◽  
In Planta ◽  
Callose Deposition ◽  
Type Iii

The type III secretion system (T3SS) of Pseudomonas syringae translocates into plant cells multiple effectors that suppress pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI). P. syringae pv. tomato DC3000 no longer delivers the T3SS translocation reporter AvrPto-Cya in Nicotiana benthamiana leaf tissue in which PTI was induced by prior inoculation with P. fluorescens(pLN18). Cosmid pLN18 expresses the T3SS system of P. syringae pv. syringae 61 but lacks the hopA1Psy61 effector gene. P. fluorescens(pLN18) expressing HrpHPtoDC3000 or HopP1PtoDC3000, two T3SS-associated putative lytic transglycosylases, suppresses PTI, based on multiple assays involving DC3000 challenge inoculum (AvrPto-Cya translocation, hypersensitive response elicitation, and colony development in planta) or on plant responses (vascular dye uptake or callose deposition). Analysis of additional mutations in pHIR11 derivatives revealed that the pLN18-encoded T3SS elicits a higher level of reactive oxygen species (ROS) than does P. fluorescens without a T3SS, that enhanced ROS production is dependent on the HrpK1 translocator, and that HopA1Psy61 suppresses ROS elicitation attributable to both the P. fluorescens PAMPs and the presence of a functional T3SS.

scholarly journals Oral sodium phenylbutyrate therapy in homozygous beta thalassemia: a clinical trial

Blood ◽  
1995 ◽  
Vol 85 (1) ◽  
pp. 43-49 ◽  
Author(s):  
AF Collins ◽  
HA Pearson ◽  
P Giardina ◽  
KT McDonagh ◽  
SW Brusilow ◽  
...  
Keyword(s):  
Fetal Hemoglobin ◽  
Cell Number ◽  
Response To Treatment ◽  
Beta Thalassemia ◽  
Thalassemia Patient ◽  
Alpha Globin ◽  
Pill Counts ◽  
Sodium Phenylbutyrate

Butyrate analogues have been shown to increase fetal hemoglobin (HbF) production in vitro and in vivo. Sodium phenylbutyrate (SPB), an oral agent used to treat individuals with urea-cycle disorders, has been shown to increase HbF in nonanemic individuals and in individuals with sickle cell disease. We have treated eleven patients with homozygous beta thalassemia (three transfusion dependent) and one sickle-beta- thalassemia patient with 20 g/d (forty 500-mg tablets) of SPB for 41 to 460 days. All patients showed an increase in the percent of F reticulocytes associated with treatment, but only four patients responded by increasing their Hb levels by greater than 1 g/dL (mean increase, 2.1 g/dL; range, 1.2 to 2.8 g/dL). None of the transfusion- dependent thalassemia subjects responded. Increase in Hb was associated with an increase in red blood cell number (mean increase, 0.62 x 10(12)/L), and mean corpuscular volume (mean increase, 6 fL). Changes in percent HbF, absolute HbF levels, or alpha- to non-alpha-globin ratios as measured by levels of mRNA and globin protein in peripheral blood did not correlate with response to treatment. Response to treatment was not associated with the type of beta-globin mutation, but baseline erythropoietin levels of greater than 120 mU/mL was seen in all responders and only two of eight nonresponders to SPB. Compliance with treatment was greater than 90% as measured by pill counts. Side effects of the drug included weight gain and/or edema caused by increase salt load in 2/12, transient epigastric discomfort in 7/12, and abnormal body odor in 3/12 subjects. Two splenectomized patients who were not on prophylactic antibiotics developed sepsis while on treatment. We conclude that SPB increases Hb in some patients with thalassemia, but the precise mechanism of action is unknown.

scholarly journals Characterization of a Tryptophan 2-Monooxygenase Gene from Puccinia graminis f. sp. tritici Involved in Auxin Biosynthesis and Rust Pathogenicity

2014 ◽  
Vol 27 (3) ◽  
pp. 227-235 ◽  
Author(s):  
Chuntao Yin ◽  
Jeong-Jin Park ◽  
David R. Gang ◽  
Scot H. Hulbert
Keyword(s):  
Pseudomonas Syringae ◽  
Rust Fungus ◽  
Infected Plant ◽  
Bacterial Pathogen ◽  
Leaf Tissue ◽  
Infected Leaf ◽  
Puccinia Graminis ◽  
Monooxygenase Gene ◽  
Microbe Interactions

The plant hormone indole-3-acetic acid (IAA) is best known as a regulator of plant growth and development but its production can also affect plant–microbe interactions. Microorganisms, including numerous plant-associated bacteria and several fungi, are also capable of producing IAA. The stem rust fungus Puccinia graminis f. sp. tritici induced wheat plants to accumulate auxin in infected leaf tissue. A gene (Pgt-IaaM) encoding a putative tryptophan 2-monooxygenase, which makes the auxin precursor indole-3-acetamide (IAM), was identified in the P. graminis f. sp. tritici genome and found to be expressed in haustoria cells in infected plant tissue. Transient silencing of the gene in infected wheat plants indicated that it was required for full pathogenicity. Expression of Pgt-IaaM in Arabidopsis caused a typical auxin expression phenotype and promoted susceptibility to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000.

Bacteria occurring in onion (Allium cepa L.) foliage in Puerto Rico.

1969 ◽  
pp. 199-219
Author(s):  
Juan Calle-Bellido ◽  
Lydia I. Rivera-Vargas ◽  
Myrna Alameda ◽  
Irma Cabrera
Keyword(s):  
Puerto Rico ◽  
Pseudomonas Syringae ◽  
Allium Cepa ◽  
Leaf Tissue ◽  
Field Conditions ◽  
Bacterial Strains ◽  
Burkholderia Glumae ◽  
Pathogenic Potential ◽  
Allium Cepa L ◽  
Leaf Tissues

Bacteria associated with foliar symptoms of onion (Allium cepa L.) were examined in the southern region of Puerto Rico from January through April 2004. Different symptoms were observed in onion foliage of cultivars 'Mercedes' and 'Excalibur' at Juana Díaz and Santa Isabel, Puerto Rico. Ellipsoidal sunken lesions with soft rot and disruption of tissue were the most common symptoms observed in onion foliage in field conditions. From a total of 39 bacterial strains isolated from diverse symptoms in onion foliage, 38% were isolated from soft rotting lesions. Ninety-two percent of the bacteria isolated from onion foliage was Gram negative. Pantoea spp. with 25%, was the most frequently isolated genus, followed by Pasteurella spp. and Serratia rubidae with 10% each. Fifty- six percent of the strains held plant pathogenic potential; these strains belong to the genera Acidovorax sp., Burkholderia sp., Clavibacter sp., Curtobacterium sp., Enterobacter sp., Pantoea spp., Pseudomonas spp., and Xanthomonas spp. Pathogenicity tests showed that seven out of eight tested bacterial strains evaluated under field conditions caused symptoms in onion foliage for both cultivars. Acidovorax avenae subsp. citrulli, Burkholderia glumae, Pantoea agglomerans, P. dispersa, Pseudomonas sp., Xanthomonas sp., and Xanthomonas-Wke sp. were pathogenic to leaf tissues. Clavibacter michiganensis was not pathogenic to leaf tissues. Other bacteria identified as associated with onion leaf tissue were Curtobacterium flaccumfaciens, Cytophaga sp., Enterobacter cloacae, Flavimonas oryzihabitans, Mannheimia haemolytica, Pantoea stewartii, Pasteurella anatis, P. bettyae, P. langaaensis, Photobacterium damselae, Pseudomonas syringae pv. aptata, Rhizobium radiobacter, Serratia rubidae, Sphingobacterium spiritivorum, Sphingomonas sanguinis, and an unknown strain. This paper is the first survey of bacteria associated with onion foliage in Puerto Rico. The role of non- phytopathogenic bacteria associated with the life cycle of onion under field conditions remains unknown.

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