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PubMedDate not supplied

[Genetic diversity in pseudomonads associated with cereal cultures infected with basal bacteriosis].

The genetic properties of 45 pseudomonad strains isolated from cereal cultures exhibiting symptoms of basal bacteriosis have been investigated. Considerable genetic diversity has been demonstrated using DNA fingerprints obtained by amplification with REP, ERIC, and BOX primers. Restriction analysis of the 16S-23S internal transcribed spacer (ITS1) allowed the strains to be subdivided into two major groups. In a phylogenetic tree, the ITS1s of these groups fell into two clusters, which also included the ITS1 of Pseudomonas syringae ("Syringae" cluster) and the ITS1 of P. fluorescens, P. tolaasii, P. reactans, P. gingeri, and P. agarici ("Fluorescens" cluster) from the GenBank database. Comparison of the ITS1 divergence levels within the "Fluorescens" cluster suggests expediency of treating P. tolaasii, P. reactans, various P. fluorescens groups, and, possibly, P. gingeri and P. agarici as subspecies of one genospecies. The intragenomic heterogeneity of ITS1s was observed in some of the pseudomonad strains studied. The results of amplification with specific primers and subsequent sequencing of the amplificate suggest the possibility of the presence of a functionally active syrB gene involved in syringomycin biosynthesis in the strains studied.

Bacterial Proteins↗
PubMedDate not supplied

[Loquat canker: a new disease for Argentina].

A stem canker disease caused by Pseudomonas syringae pv. eriobotryae (Takimoto) Young, Dye y Wilkie on loquat (Eriobotrya Japonica [Thumb] Lindl) was recorded for the first time in Argentina. Symptoms of the disease appeared as dry stem cankers which in advanced stages surrounded the stems. Similar cankers were noticeable on leaves midribs. Seven bacterial strains were isolated from diseased loquats and their identification was based on disease symptoms, pathogenicity and cultural and biochemical characteristics. All strains were levan positive and gave a hypersensitive reaction on tobacco leaves. Neither arginine dehydrolase nor oxidase was detected in any of the strains which produced a diffusible green pigment on King B which fluoresced under UV light and a distinct diffusible brown pigment on King B, SPA and Tween 80 media within 5-7 days of incubation. Lipolysis of Tween 80 was also recorded. The symptoms observed in the field and obtained by experimental inoculations were similar to those induced by Pseudomonas syringae pv. eriobotryae in the original description of the disease.

Argentina↗
PubMedDate not supplied

[The specificity of an immune serum to the exocellular lipopolysaccharide of Pseudomonas wieringae].

The serum obtained to exocellular lipopolysaccharide (ELPS) of Pseudomonas wieringae selectively agglutinated strains of pathovar of P. syringae and did not agglutinated strains of P. cichorii, P. solanacearum, P. gladioli pv. allicola, P. fluoroviolaceus, strains of nonphytopathogenic pseudomonads as well as bacteria of the genera Erwinia, Bacillus, Xanthomonas, Klebsiella. Consequently, the antigen determinant common with antigen of the species Pseudomonas syringae is present in the composition of ELPS.

Agglutination Tests↗
PubMed1987

Mucoid strains of Pseudomonas aeruginosa are devoid of mannuronan C-5 epimerase.

Mucoid strains of Azotobacter vinelandii, Pseudomonas aeruginosa and Pseudomonas syringae var glycinia synthesize alginate, an extracellular copolymer comprising D-mannuronosyl and L-guluronosyl moieties. Extracellular mannuronan C-5 epimerase, which converts polymannuronate to alginate, was demonstrated in supernatant fluid from cultures of A. vinelandii. However, the enzyme could not be demonstrated, using the same assay, in supernatant fluids of cultures of mucoid strains of P. aeruginosa or of P. syringae var glycinia, or in cell-free sonic extracts of P. aeruginosa. The results suggest that the pathways of alginate biosynthesis in A. vinelandii and Pseudomonas species may differ.

Alginates↗
PubMed2004

Functional display of foreign protein on surface of Escherichia coli using N-terminal domain of ice nucleation protein.

We investigated the ability of the N-terminal domain of InaK, an ice nucleation protein from Pseudomonas syringae KCTC1832, to act as an anchoring motif for the display of foreign proteins on the Escherichia coli cell surface. Total expression level and surface display efficiency of green fluorescent protein (GFP) was compared following their fusion with either the N-terminal domain of InaK (InaK-N), or with the known truncated InaK containing both N- and C-terminal domains (InaK-NC). We report that the InaK-N/GFP fusion protein showed a similar cell surface display efficiency ( approximately 50%) as InaK-NC/GFP, demonstrating that the InaK N-terminal region alone can direct translocation of foreign proteins to the cell surface and can be employed as a potential cell surface display motif. Moreover, InaK-N/GFP showed the highest levels of total expression and surface display based on unit cell density. InaK-N was also successful in directing cell surface display of organophosphorus hydrolase (OPH), confirming its ability to act as a display motif.

Bacterial Outer Membrane Proteins↗
PubMed2005

Bioprocess intensification in flow-through monolithic microbioreactors with immobilized bacteria.

Microporous polymers (with porosity up to 90%) with a well-prescribed internal microstructure were prepared in monolithic form to construct a flow-through microbioreactor in which phenol-degrading bacteria, Pseudomonas syringae, was immobilized. Initially, bacteria was forced seeded within the pores and subsequently allowed to proliferate followed by acclimatization and phenol degradation at various initial substrate concentrations and flow rates. Two types of microporous polymer were used as the monolithic support. These polymers differ with respect to their pore and interconnect sizes, macroscopic surface area for bacterial support, and phase volume. Polymer with a nominal pore size of 100 microm with phase volume of 90% (with highly open pore structure) yielded reduced bacterial proliferation, while the polymer with nominal pore size of 25 microm with phase volume of 85% (with small interconnect size and large pore area for bacterial adhesion) yielded monolayer bacterial proliferation. Bacteria within the 25 microm polymer support remained monolayered, without any apparent production of extracellular matrix during the 30-day continuous experimental period. The microbioreactor performance was characterized in terms of volumetric utilization rate and compared with the published data, including the case where the same bacteria was immobilized on the surface of microporous polymer beads and used in a packed bed during continuous degradation of phenol. It is shown that at similar initial substrate concentration, the volumetric utilization in the microreactor is at least 20-fold more efficient than the packed bed, depending on the flow rate of the substrate solution. The concentration of the bacteria within the pores of the microreactor decreases from 2.25 cells per microm2 on the top surface to about 0.4 cells per microm2 within 3 mm reactor depth. If the bacteria-depleted part of the microreactor is disregarded, the volumetric utilization increases by a factor of 30-fold compared with the packed bed. This efficiency increase is attributed to the reduction of diffusion path for the substrate and nutrients and enhanced availability of the bacteria for bioconversion in the absence of biofilm formation as well as the presence of flow over the surface of the monolayer bacteria.

Bioreactors↗
PubMed2006

Analysis of the defence phosphoproteome of Arabidopsis thaliana using differential mass tagging.

Despite recent advances in proteomic technologies, quantitative analysis of the proteome remains a challenging task. Phosphorylation of proteins is central to signal transduction pathways and plays an important role in plant defence against pathogens, although the immediate targets of kinases remain elusive. Determining changes in the phosphoproteome during the defence response is a major goal in molecular plant pathology. In this first description of the novel mass tagging strategy (iTRAQ Applied Biosystems) applied to plant pathogen interactions, we describe early changes to the phosphoproteome of Arabidopsis thaliana during the defence response to Pseudomonas syringae pv. tomato DC3000. We identified five proteins which showed reproducible differences between a control and three different bacterial challenges, thus identifying proteins potentially phosphorylated as part of a plant basal defence response. Four of the five proteins a dehydrin, a putative p23 co-chaperone, heat shock protein 81 and a plastid-associated protein (PAP)/fibrillin, are known to be phosphorylated or have potential phosphorylation sites. One further protein, the large subunit of Rubisco, showed a significant difference between tissue undergoing the hypersensitive response and a basal defence response. We document the reproducibility, utility and problems associated with this approach.

Arabidopsis↗
PubMed2005

Antimicrobial activity of some Ganoderma species from Nigeria.

The crude n-hexane:diethyl ether, chloroform:acetone and methanol extracts of four species of Ganoderma (Ganoderma colossum (Fr.) C. F. Baker, G. resinaceum Boud., G. lucidum (cf.) (Curtis) P. Karst. and G. boninense (cf.) Pat.), from Nigeria, were tested for antimicrobial activity. The three solvent extracts of all the species of Ganoderma were active against Pseudomonas syringae and Bacillus subtilis, whereas none of the extracts were active against Cladosporium herbarum. Preliminary thin layer chromatography chemical tests on these extracts of Ganoderma showed that they contained compounds that stained blue-violet and blue or green when sprayed with anisaldehyde-sulphuric acid or Dragendorff, respectively. The profile of compounds in the extracts showed some variation among the four species.

Anti-Bacterial Agents↗
PubMed2003

PCC1: a merging point for pathogen defence and circadian signalling in Arabidopsis.

Using a cDNA-array we identified expressed sequence tag 163B24T7 as rapidly up-regulated in Arabidopsis thaliana (L.) Heynh. after pathogen exposure. Detailed expression analysis revealed that the corresponding gene is up-regulated not only after exposure to avirulent Pseudomonas syringae pv. tomato but also to virulent strains. This up-regulation is dependent on functional salicylic acid defence-signalling pathways. Moreover, we found the gene was circadian-regulated, showing peaks of expression at the end of the day. Using plants overexpressing the clock component CCA1, we showed that the PCC1 gene is regulated by the inner clock of Arabidopsis. Accordingly, we named the gene PCC1, for pathogen and circadian controlled. PCC1 is a member of a novel family of six small polypeptides in Arabidopsis. A functional role for PCC1 in plant defence was demonstrated since plants overexpressing PCC1 are resistant against normally virulent oomycetes. Thus, PCC1 demonstrates a potential interrelationship between pathogen and circadian signalling pathways.

Amino Acid Sequence↗
PubMed2004

Light conditions influence specific defence responses in incompatible plant-pathogen interactions: uncoupling systemic resistance from salicylic acid and PR-1 accumulation.

In incompatible plant-pathogen interactions, disease resistance is generated by rapid activation of a multitude of plant defence reactions. Little is known about the dependency of these resistance responses on external factors. The plasticity of plant defence mechanisms in terms of light conditions is studied here. Interaction of Arabidopsis thaliana (L.) Heynh. with an avirulent strain of Pseudomonas syringae pv. maculicola in the dark resulted in increased apoplastic bacterial growth and therefore reduced local resistance as compared to an infection process in the presence of light. Several characteristic defence reactions, including activation of phenylalanine ammonia-lyase, accumulation of salicylic acid (SA), expression of the pathogenesis-related protein PR-1 and the development of a microscopically defined hypersensitive response, proved to be light dependent. In contrast, the extent of the oxidative burst, as estimated by induction of the protectant gene glutathione- S-transferase, was not weakened by the absence of light. Moreover, pathogen-induced accumulation of jasmonic acid, production of the phytoalexin camalexin and transcriptional induction of a pathogen-inducible myrosinase were even more pronounced in the dark. Apart from affecting local defence responses, light also influenced the establishment of systemic acquired resistance (SAR). SAR development in response to infection by avirulent bacteria was completely lost when the primary infection process occurred in the absence of light. SAR developed both under medium (70 micromol photons m(-2) s(-1)) and strong (500 micromol photons m(-2) s(-1)) light conditions but was in the latter case not associated with an accumulation of SA and PR-1 in systemic leaves, demonstrating that SAR can be executed independently from these molecular SAR markers. Our results are consistent with the notion that SA accumulation in infected primary leaves is necessary for induction of systemic resistance and indicate that defence mechanisms different from SA signalling and PR-protein action exist in systemic tissue to confer resistance during SAR.

Arabidopsis↗
PubMed2006

Identification and deletion analysis of the promoter of the pepper SAR8.2 gene activated by bacterial infection and abiotic stresses.

The pepper SAR8.2 gene, CASAR82A, was locally and systemically induced in pepper plants which had been infected by Xanthomonas campestris pv. vesicatoria or by Pseudomonas fluorescens. The DNA 1,283 bp sequence upstream of the CASAR82A gene was assessed with regard to the activity of the CASAR82A promoter fused to the beta-glucuronidase (GUS) reporter gene, via an Agrobacterium-mediated transient expression assay. In tobacco leaves which transiently expressed the -831 bp CASAR82A promoter, GUS activity was locally and systemically induced by Pseudomonas syringae pv. tabaci. GUS activity, which was driven by the -831 promoter, was also differentially activated in the leaves as the result of treatment with salicylic acid, ethylene, methyl jasmonate, abscisic acid, NaCl, and low temperatures. The -831 bp sequence upstream of the CASAR82A gene elicited full promoter activity in response to pathogen infection, abiotic elicitors, and environmental stresses. The expression of the pepper transcription factor, CARAV1, was shown to activate the CASAR82A promoter. Analyses of a series of 5'-deletions of the CASAR82A promoter revealed that novel cis-acting elements necessary for the induction of gene expression as the result of exposure to pathogen and abiotic elicitors appear to be localized in the promoter region between -831 and -759 bp.

Abscisic Acid↗
PubMed2005

Stress induces the expression of AtNADK-1, a gene encoding a NAD(H) kinase in Arabidopsis thaliana.

A novel Arabidopsis thaliana gene (AtNADK-1) was identified based on its response to radiation and oxidative stress. Levels of AtNADK-1 mRNA increase eight-fold following exposure to ionising radiation and are enhanced three-fold by treatment with hydrogen peroxide. The gene also appears to be differentially regulated during compatible and incompatible plant-pathogen interactions in response to Pseudomonas syringae pv. tomato. The full-length AtNADK-1 cDNA encodes a 58-kDa protein that shows high sequence homology to the recently defined family of NAD(H) kinases. Recombinant AtNADK-1 utilises ATP to phosphorylate both NAD and NADH, showing a two-fold preference for NADH. Using reverse genetics, we demonstrate that AtNADK-1 deficient plants display enhanced sensitivity to gamma irradiation and to paraquat-induced oxidative stress. Our results indicate that this novel NAD(H) kinase may contribute to the maintenance of redox status in Arabidopsis thaliana.

Amino Acid Sequence↗
PubMed2004

Climate change and shifts in spring phenology of three horticultural woody perennials in northeastern USA.

We evaluated spring phenology changes from 1965 to 2001 in northeastern USA utilizing a unique data set from 72 locations with genetically identical lilac plants (Syringa chinensis, clone "Red Rothomagensis"). We also utilized a previously validated lilac-honeysuckle "spring index" model to reconstruct a more complete record of first leaf date (FLD) and first flower date (FFD) for the region from historical weather data. In addition, we examined mid-bloom dates for apple (Malus domestica) and grape (Vitis vinifera) collected at several sites in the region during approximately the same time period. Almost all lilac sites with significant linear trends for FLD or FFD versus year had negative slopes (advanced development). Regression analysis of pooled data for the 72 sites indicated an advance of -0.092 day/year for FFD (P=0.003). The slope for FLD was also negative (-0.048 day/year), but not significant (P=0.234). The simulated data from the "spring index" model, which relies on local daily temperature records, indicated highly significant (P<0.001) negative slopes of -0.210 and -0.123 day/year for FLD and FFD, respectively. Data collected for apple and grape also indicated advance spring development, with slopes for mid-bloom date versus year of -0.20 day/year (P=0.01) and -0.146 (P=0.14), respectively. Collectively, these results indicate an advance in spring phenology ranging from 2 to 8 days for these woody perennials in northeastern USA for the period 1965 to 2001, qualitatively consistent with a warming trend, and consistent with phenology shifts reported for other mid- and high-latitude regions.

Climate↗
PubMed2004

The ethylene-responsive factor like protein 1 (CaERFLP1) of hot pepper (Capsicum annuum L.) interacts in vitro with both GCC and DRE/CRT sequences with different binding affinities: possible biological roles of CaERFLP1 in response to pathogen infection and high salinity conditions in transgenic tobacco plants.

From a pathogen-inoculated hot pepper (Capsicum annuum L. cv. Pukang) leaf EST, we identified a cDNA clone, pCaERFLP1, encoding a putative transcription factor that contains a single ERF/AP2 DNA binding domain. CaERFLP1 was most closely related to tomato LeERF2 (73%), both of which belong to the novel ERF class IV typified by the N-terminal MCGGAIL signature sequence, while it had a limited sequence identity (25-30%) with Arabidopsis AtERFs and tobacco NtERFs. Quantitative gel retardation assays revealed that bacterially expressed full-length CaERFLP1 was able to form a specific complex with both the GCC box and DRE/CRT motif, with its binding affinity for GCC being stronger than for DRE/CRT. When fused to the GAL4 DNA binding domain, the N-terminal CaERFLP1(1-37) and C-terminal CaERFLP1(198-264) mutant polypeptides could function individually as transactivators in yeast. This suggests that two separate domains of CaERFLP1 may play distinct roles in transcription activation. In particle co-bombardment experiments, CaERFLP1 activated the transcription of reporter genes containing the 4X[GCC] element in tobacco cells. In hot pepper plants, the steady-state level of CaERFLP1 mRNA was markedly induced by multiple environmental factors, such as pathogen infection, ethylene, mechanical wounding and high salinity. Furthermore, ectopic expression of CaERFLP1 in transgenic tobacco plants resulted in partially improved tolerance against the bacterial pathogen Pseudomonas syringae and salt stress (100 mM NaCl). Consistently, various defense-related genes, including GCC box-containing PR genes and the DRE/CRT-containing LTI45 (ERD10) gene, were constitutively expressed in 35S::CaERFLP1 tobacco plants. Thus, it appears that CaERFLP1 is functional in tobacco cells, where it induces the transactivation of some GCC- and DRE/CRT-genes to trigger a subset of stress response. Here, the possible biological role(s) of CaERFLP1 is discussed, especially with regard to the possibility that CaERFLP1 has multiple functions in the regulation of GCC- and DRE/CRT-mediated gene expression in hot pepper plants.

Amino Acid Sequence↗
PubMed2004

Overexpression of the AP2/EREBP transcription factor OPBP1 enhances disease resistance and salt tolerance in tobacco.

Osmotin promoter binding protein 1 (OPBP1), an AP2/EREBP-like transcription factor of tobacco (Nicotiana tabacum), was isolated using a yeast one-hybrid system. RNA gel blot analysis indicated that expression of the OPBP1 gene was induced by elicitor cryptogein, NaCl, ethephon, methyl jasmonate, as well as cycloheximide. Transient expression analysis using an OPBP1-eGFP fusion gene in onion epidermal cells revealed that the OPBP1 protein was targeted to the nuclear. Further, electrophoretic mobility shift assays demonstrated that the recombinant OPBP1 protein could bind to an oligonucleotide containing the GCC-box cis element. Transgenic tobacco plants with an over expression of the OPBP1 gene accumulated high levels of PR-1a and PR-5d genes and exhibited enhanced resistance to infection by Pseudomonas syringae pv tabaci and Phytophthora parasitica var nicotianae pathogens. They also exhibited increased tolerance to salt stress. These results suggest that OPBP1 might be a transcriptional regulator capable of regulating expression in sets of stress-related genes.

Acetates↗
PubMed2004

CAZFP1, Cys2/His2-type zinc-finger transcription factor gene functions as a pathogen-induced early-defense gene in Capsicum annuum.

A pepper zinc-finger protein gene, CAZFP1 , encoding the Cys2/His2-type zinc-finger transcription factor was isolated from pepper leaves inoculated with an avirulent strain Bv5-4a of Xanthomonas campestris pv. vesicatoria . The CAZFP1 protein is a nuclear targeting protein, which functions as a transcriptional regulator. The full-length CAZFP1 had no transcriptional activation activity, whereas the C-terminal region of CAZFP1 had transactivation activity. The CAZFP1 transcripts were constitutively expressed in the pepper stem, root, flower and red fruit, but were not detectable in the leaf and green fruit. The CAZFP1 transcripts accumulated earlier than the CAZFP1 (PR-1) gene in the incompatible interaction of the pepper leaves with X. campestris pv. vesicatoria . The CAZFP1 transcripts were significantly induced in the systemic, uninoculated leaf tissues early after inoculation with bacterial pathogens, but gradually declined thereafter. The CAZFP1 transcripts were localized, and confined to the phloem cells of the vascular bundle in the pepper leaf midrib in response to Colletotrichum. coccodes infection, ethylene and abscisic acid. The CAZFP1 gene was also induced much earlier by abiotic elicitors and environmental stresses, compared with the CAZFP1 gene. Overexpression of the CAZFP1 gene in the transgenic Arabidopsis plants enhanced not only the resistance against infection by Pseudomonas syringae pv. tomato, but also the drought tolerance. These results suggest that the CAZFP1 gene functions as an early-defense gene to enhance disease resistance and drought tolerance.

Amino Acid Sequence↗
PubMed2005

A hypersensitive response-induced ATPase associated with various cellular activities (AAA) protein from tobacco plants.

The hypersensitive response (HR) is one of the most critical defense systems in higher plants. In order to understand its molecular basis, we have screened tobacco genes that are transcriptionally activated during the early stage of the HR by the differential display method. Among six genes initially identified, one was found encoding a 57 kDa polypeptide with 497 amino acids not showing significant similarity to any reported proteins except for the AAA domain (ATPase associated with various cellular activities) spanning over 230 amino acids. The bacterially expressed protein exhibited ATP hydrolysis activity, and a green fluorescent protein-fusion protein localized in the cytoplasm of onion epidermis cells. The protein was subsequently designated as NtAAA1 (Nicotiana tabacum AAA1). NtAAA1 transcripts were induced 6 h after HR onset not only by TMV but also by incompatible Psuedomonas syringae, indicating that NtAAA1 is under the control of the N-gene with a common role in pathogen responses. Expression of NtAAA1 was induced by jasmonic acid and ethylene, but not by salicylic acid (SA). It also occurred at a high level in SA-deficient tobacco plants upon TMV infection. When NtAAA1 was silenced by the RNAi method, accumulation of transcripts for PR-1a significantly increased during the HR. Treatments with SA induced higher expression of PR-1a and acidic PR-2 in RNAi transgenic plants than in wild-type counterparts. These results suggest that NtAAA1 mitigates the SA signaling pathway, and therefore that NtAAA1 modulates the pathogen response of the host plants by adjusting the HR to an appropriate level.

Adenosine Triphosphatases↗
PubMed2005

Colonization of the Arabidopsis rhizosphere by fluorescent Pseudomonas spp. activates a root-specific, ethylene-responsive PR-5 gene in the vascular bundle.

Plants of which the roots are colonized by selected strains of non-pathogenic, fluorescent Pseudomonas spp. develop an enhanced defensive capacity against a broad spectrum of foliar pathogens. In Arabidopsis thaliana, this rhizobacteria-induced systemic resistance (ISR) functions independently of salicylic acid but requires responsiveness to jasmonic acid and ethylene. In contrast to pathogen-induced systemic acquired resistance (SAR), ISR is not associated with systemic changes in the expression of genes encoding pathogenesis-related (PR) proteins. To identify genes that are specifically expressed in response to colonization of the roots by ISR-inducing Pseudomonas fluorescens WCS417r bacteria, we screened a collection of Arabidopsis enhancer trap and gene trap lines containing a transposable element of the Ac/Ds system and the GUS reporter gene. We identified an enhancer trap line (WET121) that specifically showed GUS activity in the root vascular bundle upon colonization of the roots by WCS417r. Fluorescent Pseudomonas spp. strains P. fluorescens WCS374r and P. putida WCS358r triggered a similar expression pattern, whereas ISR-non-inducing Escherichia coli bacteria did not. Exogenous application of the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC) mimicked the rhizobacteria-induced GUS expression pattern in the root vascular bundle, whereas methyl jasmonic acid and salicylic acid did not, indicating that the Ds element in WET121 is inserted in the vicinity of an ethylene-responsive gene. Analysis of the expression of the genes in the close vicinity of the Ds element revealed AtTLP1 as the gene responsible for the in cis activation of the GUS reporter gene in the root vascular bundle. AtTLP1 encodes a thaumatin-like protein that belongs to the PR-5 family of PR proteins, some of which possess antimicrobial properties. AtTLP1 knockout mutant plants showed normal levels of WCS417r-mediated ISR against the bacterial leaf pathogen Pseudomonas syringae pv. tomato DC3000, suggesting that expression of AtTLP1 in the roots is not required for systemic expression of ISR in the leaves. Together, these results indicate that induction of AtTLP1 is a local response of Arabidopsis roots to colonization by non-pathogenic fluorescent Pseudomonas spp. and is unlikely to play a role in systemic resistance.

Amino Acid Sequence↗