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PubMed2004

Polysaccharides of pseudomonas pathovar strains that infect pea, tomato, and soya bean.

In order to understand the mode of action of taxonomically related Pseudomonas syringae pathovar strains that infect pea, tomato, and soya bean, we examined their extracellular polysaccharides (EPS). Maximum production of polysaccharide in shake culture of these pathogens was observed between 24 and 60 h. P. syringae pv. pisi 519, the bacterial blight pathogen of pea, produced a higher amount of polysaccharide (34.87 microg/mL) at 60 h compared with 32.67 microg/mL produced by P. syringae pv. glycinea NCPPB 1783, the bacterial blight pathogen of soya bean, and 30.03 microg/mL produced by P. syringae pv. tomato NCPPB 269, the bacterial speck pathogen of tomato. EPS produced by P. syringae pv. pisi 519, P. syringae pv. tomato NCPPB 269, and P. syringae pv. glycinea NCPPB 1783 was characterized with infrared (FTIR), nuclear magnetic resonance (NMR), high performance thin layer chromatography, (HPTLC), and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. HPTLC profiles revealed the presence of glucose and glucuronic acid in all bacteria and mannose only in P. syringae pv. tomato. Molecular mass of EPS of P. syringae pv. pisi (m/z 933.8), P. syringae pv. tomato (m/z 950.4), and P. syringae pv. glycinea (m/z 933.5) was confirmed by MALDI-TOF mass spectrometry.

Fabaceae↗
PubMed1995

Development of a microbial community of bacterial and yeast antagonists to control wound-invading postharvest pathogens of fruits.

Two antagonists, the bacterium Pseudomonas syringae and the pink yeast Sporobolomyces roseus, against blue mold (caused by Penicillium expansum) on apple controlled this disease more effectively when combined at approximately equal biomass (50:50 of the same turbidity) than in individual applications. Addition of L-asparagine enhanced the biocontrol effectiveness of P. syringae but decreased that of S. roseus and had no significant effect when the antagonists were combined. Populations of both antagonists increased in apple wounds and were further stimulated by the addition of L-asparagine. The carrying capacity of wounds for P. syringae was not affected by S. roseus. Populations of P. syringae in wounds inoculated individually or in a 50:50 mixture with S. roseus reached the same level after 3 days at 22 degrees C. However, populations of S. roseus recovered after applications of the mixture were consistently lower than those recovered after individual applications. Similar effects were observed in in vitro tests in which populations of S. roseus grown in mixtures with P. syringae were consistently lower than those grown alone, while the populations of P. syringae were not affected by the presence of S. roseus. A total of 36 carbon and 35 nitrogen compounds were tested for utilization by both antagonists. Fourteen nitrogenous compounds were utilized by both P. syringae and S. roseus, and an additional nine compounds were utilized by P. syringae. S. roseus and P. syringae utilized 17 and 13 carbon sources, respectively; 9 sources were common to both antagonists. Populations of these antagonists in apple wounds appear to form a relatively stable community dominated by P. syringae.(ABSTRACT TRUNCATED AT 250 WORDS)

Biomass↗
PubMed2006

Impact of botanical pesticides derived from Melia azedarach and Azadirachta indica plants on the emission of volatiles that attract Parasitoids of the diamondback moth to cabbage plants.

Herbivorous and carnivorous arthropods use chemical information from plants during foraging. Aqueous leaf extracts from the syringa tree Melia azedarach and commercial formulations from the neem tree Azadirachta indica, Neemix 4.5, were investigated for their impact on the flight response of two parasitoids, Cotesia plutellae and Diadromus collaris. Cotesia plutellae was attracted only to Plutella xylostella-infested cabbage plants in a wind tunnel after an oviposition experience. Female C. plutellae did not distinguish between P. xylostella-infested cabbage plants treated with neem and control P. xylostella-infested plants. However, females preferred infested cabbage plants that had been treated with syringa extract to control infested plants. Syringa extract on filter paper did not attract C. plutellae. This suggests that an interaction between the plant and the syringa extract enhances parasitoid attraction. Diadromus collaris was not attracted to cabbage plants in a wind tunnel and did not distinguish between caterpillar-damaged and undamaged cabbage plants. Headspace analysis revealed 49 compounds in both control cabbage plants and cabbage plants that had been treated with the syringa extract. Among these are alcohols, aldehydes, ketones, esters, terpenoids, sulfides, and an isothiocyanate. Cabbage plants that had been treated with the syringa extract emitted larger quantities of volatiles, and these increased quantities were not derived from the syringa extract. Therefore, the syringa extract seemed to induce the emission of cabbage volatiles. To our knowledge, this is the first example of a plant extract inducing the emission of plant volatiles in another plant. This interesting phenomenon likely explains the preference of C. plutellae parasitoids for cabbage plants that have been treated with syringa extracts.

Animals↗
PubMed2004

Study of the effect of Ammoides pusilla (Brot.) Breist, essential oil against Pseudomonas sp.

The economic losses caused by plant pathogens, the development of genetic resistance to applied pesticides by major pathogenic bacteria and public concerns about the use of synthetic chemicals on food and environment, increased interest in the search of alternative safe methods to control diseases spreading. Biological control with plant extracts has emerged as a promising option. The present study was carried out to determine the effect of Ammoides pusilla essential oil on the growth of two pathogenic bacteria, Pseudomonas syringae pv. syringae and Pseudomonas syringae pv. mosprunorum Three essential oil treatments, 1/2 v/v, 1/5 v/v and 1/10 v/v concentrations, were tested onto these bacteria, using the agar diffusion test. The results showed that 1/2 v/v concentration treatment completely inhibited Pseudomonas syringae pv. mosprunorum growth and produced a 35 mm diameter inhibition zone for Pseudomonas syringae pu. syringae. 1/5 v/v essential oil concentration produced inhibition zones of 19 and 17 mm diameter for Pseudomonas syringae pv. syringae and Pseudormonas syringae pv. mosprunorum respetively, while 1/10 v/v essential oil concentration gave inhibition plates of 11 and 12 mm diameter for both stains in the order cited. In in vitro culture essential oil gave promising results suggesting that future studies should be carried out and focus on determining appropriate formulation of this bio-molecule to be use as a bio-control agent.

Algeria↗
PubMed1984

Differences between lipopolysaccharide compositions of plant pathogenic and saprophytic pseudomonas species.

Lipopolysaccharides (LPS) were obtained by washing cells of plant pathogenic and saprophytic Pseudomonas species with saline (fraction 1) and then with saline-EDTA (fraction 2). The cells subsequently were extracted with phenol to yield a third aqueous preparation (fraction 3). Each fraction type contained the LPS components, lipid A, heptose, 2-keto-3-deoxy sugar, and neutral and amino sugars. The neutral sugar compositions of fractions 1, 2, and 3, although similar within a species, differed between the Pseudomonas species. The LPS of two pathovars (pv.) of Pseudomonas syringae had glucose and rhamnose as major components: 13 (+/-3)% glucose and 87 (+/-3)% rhamnose for P. syringae pv. pisi and 18 (+/-5)% glucose and 76 (+/-2)% rhamnose for P. syringae pv. syringae. Fucose was present in addition to glucose and rhamnose for P. syringae pv. phaseolicola (68 [+/-8]% rhamnose, 14 [+/-1]% fucose, and 14 [+/-5]% glucose) and P. syringae pv. tabaci (24 [+/-2]% rhamnose, 54 [+/-3]% fucose, and 17 [+/-1]% glucose). The LPS from different races of P. syringae pv. pisi and P. syringae pv. phaseolicola could not be distinguished by neutral sugar composition. Three saprophytic species, P. aeruginosa, P. fluorescens, and P. putida, also produced LPS which had different proportions of rhamnose, fucose, and glucose. The LPS from three isolates of P. putida were distinct in possessing a high proportion of amino sugar and containing glucose as the major neutral sugar component (86 to 100%). The LPS fractions from plant pathogenic and saprophytic Pseudomonas species did not elicit browning or phytoalexin production in treated dark red kidney bean cotyledons or red Mexican bean leaves. Rather, chlorosis of the LPS-treated leaf tissue was observed.

Journal Article↗
PubMed1994

Cloning and expression of genes required for coronamic Acid (2-ethyl-1-aminocyclopropane 1-carboxylic Acid), an intermediate in the biosynthesis of the phytotoxin coronatine.

Coronamic acid (CMA; 2-ethyl-1-aminocyclopropane 1-carboxylic acid) is an intermediate in the biosynthesis of coronatine (COR), a chlorosis-inducing phytotoxin produced by Pseudomonas syringae pv. glycinea PG4180. Tn5 mutagenesis and substrate feeding studies were previously used to characterize regions of the COR biosynthetic gene cluster required for synthesis of coronafacic acid and CMA, which are the only two characterized intermediates in the COR biosynthetic pathway. In the present study, additional Tn5 insertions were generated to more precisely define the region required for CMA biosynthesis. A new analytical method for CMA detection which involves derivatization with phenylisothiocyanate and detection by high-performance liquid chromatography (HPLC) was developed. This method was used to analyze and quantify the production of CMA by selected derivatives of P. syringae pv. glycinea which contained mutagenized or cloned regions from the CMA biosynthetic region. pMU2, a clone containing a 6.45-kb insert from the CMA region, genetically complemented mutants which required CMA for COR production. When pMU2 was introduced into P. syringae pv. glycinea 18a/90 (a strain which does not synthesize COR or its intermediates), CMA was not produced, indicating that pMU2 does not contain the complete CMA biosynthetic gene cluster. However, when two plasmid constructs designated pMU234 (12.5 kb) and pKTX30 (3.0 kb) were cointroduced into 18a/90, CMA was detected in culture supernatants by thin-layer chromatography and HPLC. The biological activity of the CMA produced by P. syringae pv. glycinea 18a/90 derivatives was demonstrated by the production of COR in cosynthesis experiments in which 18a/90 transconjugants were cocultivated with CMA-requiring mutants of P. syringae pv. glycinea PG4180. CMA production was also obtained when pMU234 and pKTX30 were cointroduced into P. syringae pv. syringae B1; however, these two constructs did not enable Escherichia coli K-12 to synthesize CMA. The production of CMA in P. syringae strains which lack the COR biosynthetic gene cluster indicates that CMA production can occur independently of coronafacic acid biosynthesis and raises interesting questions regarding the evolutionary origin of the COR biosynthetic pathway.

Journal Article↗
PubMed1992

The cloned avirulence gene avrPto induces disease resistance in tomato cultivars containing the Pto resistance gene.

Resistance of tomato plants to the bacterial pathogen Pseudomonas syringae pv. tomato race 0 is controlled by the locus Pto. A bacterial avirulence gene was cloned by constructing a cosmid library from an avirulent P. syringae pv. tomato race, conjugating the recombinants into a strain of P. syringae pv. maculicola virulent on a tomato cultivar containing Pto, and screening for those clones that converted the normally virulent phenotype to avirulence. The cloned gene, designated avrPto, reduced multiplication of P. syringae pv. tomato transconjugants specifically on Pto tomato lines, as demonstrated by bacterial growth curve analyses. Analysis of F2 populations revealed cosegregation of resistance to P. syringae pv. tomato transconjugants carrying avrPto with resistance to P. syringae pv. tomato race 0. Surprisingly, mutation of avrPto in P. syringae pv. tomato race 0 does not eliminate the avirulent phenotype of race 0, suggesting that additional, as yet uncharacterized, avirulence genes and/or resistance genes may contribute to specificity in the avrPto-Pto interaction. Genetic analysis indicates that this resistance gene(s) would be tightly linked to Pto. Interestingly, P. syringae pv. glycinea transconjugants carrying avrPto elicit a typical hypersensitive resistant response in the soybean cultivar Centennial, suggesting conservation of Pto function between two crop plants, tomato and soybean.

Cell Division↗
PubMed2006

Insight into Types I and II nonhost resistance using expression patterns of defense-related genes in tobacco.

Plants protect themselves against pathogens using a range of response mechanisms. There are two categories of nonhost resistance: Type I, which does not result in visible cell death; and Type II, which entails localized programmed cell death (or hypersensitive response) in response to nonhost pathogens. The genes responsible for these two systems have not yet been intensively investigated at the molecular level. Using tobacco plants (Nicotiana tabacum), we compared expression of 12 defense-related genes between a Type I (Xanthomonas axonopodis pv. glycines 8ra) nonhost interaction, and two Type II (Pseudomonas syringae pv. syringae 61 and P. syringae pv. phaseolicola NPS3121) nonhost interactions, as well as those expressed during R gene-mediated resistance to Tobacco mosaic virus. In general, expression of most defense-related genes during R gene-mediated resistance was activated 48 h after challenge by TMV; the same genes were upregulated as early as 9 h after infiltration by nonhost pathogens. Surprisingly, X. axonopodis pv. glycines (Type I) elicited the same set of defense-related genes as did two pathovars of P. syringae, despite the absence of visible cell death. In two examples of Type II nonhost interactions, P. syringae pv. phaseolicola NPS3121 produced an expression profile more closely resembling that of X. axonopodis pv. glycines 8ra, than that of P. syringae pv. syringae 61. These results suggest that Type I nonhost resistance may act as a mechanism providing a more specific and active defense response against a broad range of potential pathogens.

Cell Death↗
PubMed1998

Preliminary description of biocidal (syringomycin) activity in fluorescent plant pathogenic Pseudomonas species.

Strains representing the fluorescent plant pathogenic Pseudomonas spp., Ps. agarici, Ps. asplenii, Ps. avellanae, Ps. beteli, Ps. caricapapayae, Ps. cichorii, Ps. corrugata, Ps. ficuserectae, Ps. flectens, Ps. fuscovaginae, Ps. marginalis, Ps. meliae, Ps. savastanoi, Ps. syringae, Ps. tolaasii and Ps. viridiflava were tested for biocidal activity using Aspergillus niger as assay organism. Inhibitory behaviour was found in strains of Ps. asplenii, Ps. blatchfordae, Ps. cichorii, Ps. corrugata, Ps. fuscovaginae, Ps. marginalis, Ps. marginalis pv. pastinacea, Ps. syringae pv. syringae, Ps. syringae pv. aptata, Ps. syringae pv. atrofaciens, Ps. syringae pv. lapsa, Ps. tolaasii, and strains of a Pseudomonas sp. pathogenic to Actinidia, in the Ps. savastanoi genomic sp. Antifungal activity could be identified with the production of members of the syringomycin family of toxins by strains in Ps. syringae, Ps. asplenii and Ps. fuscovaginae. These toxin reactions support suggestions made elsewhere of the synonym of the latter two species. In a preliminary characterization using tests for stability to heat, protease, acid and alkaline treatments, unknown toxins consistent with syringomycin-like toxins the strains from Actinidia species. The toxins from Ps. cichorii and from Ps. corrugata differed in their reactions from all other agents. Pseudomonas tolaasii produces the antifungal compound tolaasin. The white line reaction with Ps. reactions, a test for tolaasin production by strains of Ps. tolaasii, was confirmed as specific for this compound. Some of these low molecular weight toxins may be produced by some of these plant pathogenic strains.

Antifungal Agents↗
PubMed1986

Effects of Tabtoxinine-beta-Lactam on Nitrogen Metabolism in Avena sativa L. Roots.

The effects of tabtoxinine-beta-lactam (T-beta-L) on nitrate uptake and glutamine synthetase (GS) and nitrate reductase (NR) activities in roots of Avena sativa seedlings were determined. Seven-day-old oat seedlings placed in a 10 mm KNO(3) and 0.5 mm T-beta-L solution for 24 hours took up T-beta-L and lost approximately 90% of their root GS activity. [(3)H]-T-beta-L taken up by roots of seven-day-old oat seedlings was associated with GS immunoprecipitated from the extract of these roots. Total nitrate uptake and in vivo NR activity were decreased approximately 50% in the T-beta-L treated roots. However, T-beta-L uptake did not affect the induction phases of nitrate uptake or reduction, nor did it inhibit in vitro NR activity. Thus, the decrease in nitrate uptake and reduction is a secondary effect of T-beta-L action. Roots of seven-day-old oat seedlings were inoculated with Pseudomonas syringae pv tabaci (Tox+) and the pathogen population in the rhizosphere was estimated by dilution plate count; 6 x 10(13) bacteria were recovered after 3 days, as compared to the original inoculation with 7 x 10(9) bacteria, indicating a significant growth of the pathogen in the rhizosphere. The bacteria recovered from the rhizosphere caused chlorosis in tobacco leaves and produced T-beta-L in culture; 1 x 10(14) bacteria were recovered from roots of seedlings inoculated with P. syringae pv tabaci (Tox-) using the same inoculation and assay procedure as for the pv tabaci (Tox+). Extracts of surface-sterilized roots previously inoculated with P. syringae pv tabaci (Tox+) did not produce viable bacterial cultures when plated out on a complete medium. Oat seedlings growing in sand culture and inoculated with P. syringae pv tabaci (Tox+) had developed chlorosis, and root GS activity had declined to less than 10% of controls after 3 days. Conversely, seedlings inoculated with P. syringae pv tabaci (Tox-) never developed chlorosis and maintained normal levels of GS activity. All oat plants inoculated with P. syringae pv tabaci (Tox+) died within 7 days after inoculation as compared to the plants inoculated with P. syringae pv tabaci (Tox-) which grew to maturity.

Journal Article↗
PubMed1994

Coexistence among Epiphytic Bacterial Populations Mediated through Nutritional Resource Partitioning.

The levels of coexistence between Pseudomonas syringae and various nonpathogenic epiphytic species in the phyllosphere of beans (Phaseolus vulgaris) were assessed by using replacement series. The epiphytic species Pseudomonas fluorescens, Pantoea agglomerans, Stenotrophomonas maltophilia, and Methylobacterium organophilum were all capable of exhibiting higher levels of coexistence with P. syringae than was observed with a near-isogenic P. syringae strain pair. The ecological similarity of the epiphytes was estimated with niche overlap indices derived from in vitro carbon source utilization profiles. The level of coexistence of the epiphytes was inversely correlated with the ecological similarity of the strains. Hence, the level of coexistence between the epiphytes was proportional to the degree of niche differentiation, defined as the ability to utilize carbon sources not utilized by a competing strain. Comparisons of utilization profiles for groups of carbon sources (amino acids, organic acids, and carbohydrates) indicated the types of carbon sources for which the strains likely competed in the bean phyllosphere. P. fluorescens and P. syringae strains probably competed for most carbon sources. S. maltophilia and M. organophilum strains probably competed with P. syringae for most organic acids but few amino acids or carbohydrates. P. agglomerans strains probably competed with P. syringae for most amino acids and organic acids but few carbohydrates. A variable level of coexistence observed between P. agglomerans and P. syringae probably reflected the variability in abundance in the bean phyllosphere of the carbohydrates that P. agglomerans utilized exclusively.

Journal Article↗
PubMed2001

Acyl-homoserine lactone production is more common among plant-associated Pseudomonas spp. than among soilborne Pseudomonas spp.

A total of 137 soilborne and plant-associated bacterial strains belonging to different Pseudomonas species were tested for their ability to synthesize N-acyl-homoserine lactones (NAHL). Fifty-four strains synthesized NAHL. Interestingly, NAHL production appears to be more common among plant-associated than among soilborne Pseudomonas spp. Indeed, 40% of the analyzed Pseudomonas syringae strains produced NAHL which were identified most often as the short-chain NAHL, N-hexanoyl-L-homoserine lactone, N-(3-oxo-hexanoyl)-homoserine lactone, and N-(3-oxo-octanoyl)-L-homoserine lactone (no absolute correlation between genomospecies of P. syringae and their ability to produce NAHL could be found). Six strains of fluorescent pseudomonads, belonging to the species P. chlororaphis, P. fluorescens, and P. putida, isolated from the plant rhizosphere produced different types of NAHL. In contrast, none of the strains isolated from soil samples were shown to produce NAHL. The gene encoding the NAHL synthase in P. syringae pv. maculicola was isolated by complementation of an NAHL-deficient Chromobacterium mutant. Sequence analysis revealed the existence of a luxI homologue that we named psmI. This gene is sufficient to confer NAHL synthesis upon its bacterial host and has strong homology to psyI and ahlI, two genes involved in NAHL production in P. syringae pv. tabaci and P. syringae pv. syringae, respectively. We identified another open reading frame that we termed psmR, transcribed convergently in relation to psmI and partly overlapping psmI; this gene encodes a putative LuxR regulatory protein. This gene organization, with luxI and luxR homologues facing each other and overlapping, has been found so far only in the enteric bacteria Erwinia and Pantoea and in the related species P. syringae pv. tabaci.

4-Butyrolactone↗
PubMed1998

Characterization of the rice pathogen-related protein Rir1a and regulation of the corresponding gene.

In rice (Oryza sativa L.), local acquired resistance against Pyricularia oryzae (Cav.), the causal agent of rice blast, can be induced by a preinoculation with the non-host pathogen Pseudomonas syringae pv. syringae. We have cloned a cDNA (Rir1a) and a closely related gene (Rir1b) corresponding to transcripts that accumulate in leaf tissue upon inoculation with P. syringae pv. syringae. The cDNA encodes a putative 107 amino acid protein, Rir1a, that exhibits a putative signal peptide cleavage site in its hydrophobic N-terminal part and a C-terminal part that is relatively rich in glycine and proline. The Rir1b gene contains a Tourist and a Wanderer miniature transposable element in its single intron and encodes a nearly identical protein. Rir1a is similar in sequence (ca. 35% identical and ca. 60% conservatively changed amino acids) to the putative Wir1 family of proteins that are encoded by pathogen-induced transcripts in wheat. Using antibodies raised against a Rir1a-fusion protein we show that Rir1a is secreted from rice protoplasts transiently expressing a 35S::Rir1a construct and that the protein accumulates in the cell wall compartment of rice leaves upon inoculation with P. syringae pv. syringae. Possible roles of Rir1a in pathogen defense are discussed.

Amino Acid Sequence↗
PubMed1991

Homologous Streptomycin Resistance Gene Present among Diverse Gram-Negative Bacteria in New York State Apple Orchards.

The streptomycin resistance gene of Pseudomonas syringae pv. papulans Psp36 was cloned into Escherichia coli and used to develop a 500-bp DNA probe that is specific for streptomycin resistance in P. syringae pv. papulans. The probe is a portion of a 1-kb region shared by three different DNA clones of the resistance gene. In Southern hybridizations, the probe hybridized only with DNA isolated from streptomycin-resistant strains of P. syringae pv. papulans and not with the DNA of streptomycin-sensitive strains. Transposon insertions within the region of DNA shared by the three clones resulted in loss of resistance to streptomycin. Colony hybridization of bacteria isolated from apple leaves and orchard soil indicated that 39% of 398 streptomycin-resistant bacteria contained DNA that hybridized to the probe. These included all strains of P. syringae pv. papulans and some other fluorescent pseudomonads and nonfluorescent gram-negative bacteria, but none of the gram-positive bacteria. The same-size restriction fragments hybridized to the probe in P. syringae pv. papulans. Restriction fragment length polymorphism of this region was occasionally observed in strains of other taxonomic groups of bacteria. In bacteria other than P. syringae pv. papulans, the streptomycin resistance probe hybridized to different-sized plasmids and no relationship between plasmid size and taxonomic group or between plasmid size and orchard type, soil association, or leaf association could be detected.

Journal Article↗
PubMed1999

Location and survival of leaf-associated bacteria in relation to pathogenicity and potential for growth within the leaf

The growth and survival of pathogenic and nonpathogenic Pseudomonas syringae strains and of the nonpathogenic species Pantoea agglomerans, Stenotrophomonas maltophilia, and Methylobacterium organophilum were compared in the phyllosphere of bean. In general, the plant pathogens survived better than the nonpathogens on leaves under environmental stress. The sizes of the total leaf-associated populations of the pathogenic P. syringae strains were greater than the sizes of the total leaf-associated populations of the nonpathogens under dry conditions but not under moist conditions. In these studies the surface sterilants hydrogen peroxide and UV irradiation were used to differentiate cells that were fully exposed on the surface from nonexposed cells that were in "protected sites" that were inaccessible to these agents. In general, the population sizes in protected sites increased with time after inoculation of plants. The proportion of bacteria on leaves that were in protected sites was generally greater for pathogens than for nonpathogens and was greater under dry conditions than under moist conditions. When organisms were vacuum infiltrated into leaves, the sizes of the nonexposed "internal" populations were greater for pathogenic P. syringae strains than for nonpathogenic P. syringae strains. The sizes of the populations of the nonpathogenic species failed to increase or even decreased. The sizes of nonexposed populations following spray inoculation were correlated with the sizes of nonexposed, internal populations which developed after vacuum infiltration and incubation. While the sizes of the populations of the pathogenic P. syringae strains increased on leaves under dry conditions, the sizes of the populations of the nonpathogenic strains of P. syringae, P. agglomerans, and S. maltophilia decreased when the organisms were applied to plants. The sizes of the populations on dry leaves were also correlated with the sizes of the nonexposed populations that developed following vacuum infiltration. Although pathogenicity was not required for growth in the phyllosphere under high-relative-humidity conditions, pathogenicity apparently was involved in the ability to access and/or multiply in certain protected sites in the phyllosphere and in growth on dry leaves.

Journal Article↗
PubMed2004

Occurrence of the strA-strB streptomycin resistance genes in Pseudomonas species isolated from kiwifruit plants.

The occurrence of strA-strB streptomycin-resistance genes within transposon Tn5393 was examined in Pseudomonas syringae pv. actinidiae, P. syringae pv. syringae, and P. marginalis, isolated from kiwifruit plants in Korea and Japan. PCR amplification with primers specific to strA-strB revealed that three of the tested Pseudomonas species harbored these genes for a streptomycin-resistance determinant. Tn5393, containing strA-strB, was also identified with PCR primers designed to amplify parts of tnpA, res, and tnpR. No IS elements were detected within tnpR, nor were they found in the intergenic region between tnpR and strA. Nucleotide sequence analysis indicated that the strA sequence of P. syringae pv. actinidiae contained a single nucleotide alteration at position 593 (CAA-->CGA), as compared to Tn5393a in P. syringae pv. syringae. This resulted in an amino acid change, from Gln to Arg.

Actinidia↗
PubMed2004

Characterization of CmaA, an adenylation-thiolation didomain enzyme involved in the biosynthesis of coronatine.

Several pathovars of Pseudomonas syringae produce the phytotoxin coronatine (COR), which contains an unusual amino acid, the 1-amino-2-ethylcyclopropane carboxylic acid called coronamic acid (CMA), which is covalently linked to a polyketide-derived carboxylic acid, coronafacic acid, by an amide bond. The region of the COR biosynthetic gene cluster proposed to be responsible for CMA biosynthesis was resequenced, and errors in previously deposited cmaA sequences were corrected. These efforts allowed overproduction of P. syringae pv. glycinea PG4180 CmaA in P. syringae pv. syringae FF5 as a FLAG-tagged protein and overproduction of P. syringae pv. tomato CmaA in Escherichia coli as a His-tagged protein; both proteins were in an enzymatically active form. Sequence analysis of CmaA indicated that there were two domains, an adenylation domain (A domain) and a thiolation domain (T domain). ATP-(32)PP(i) exchange assays showed that the A domain of CmaA catalyzes the conversion of branched-chain L-amino acids and ATP into the corresponding aminoacyl-AMP derivatives, with a kinetic preference for L-allo-isoleucine. Additional experiments demonstrated that the T domain of CmaA, which is posttranslationally modified with a 4'-phosphopantetheinyl group, reacts with the AMP derivative of L-allo-isoleucine to produce an aminoacyl thiolester intermediate. This covalent species was detected by incubating CmaA with ATP and L-[G-(3)H]allo-isoleucine, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis. It is postulated that the L-allo-isoleucine covalently tethered to CmaA serves as the substrate for additional enzymes in the CMA biosynthetic pathway that catalyze cyclopropane ring formation, which is followed by thiolester hydrolysis, yielding free CMA. The availability of catalytically active CmaA should facilitate elucidation of the details of the subsequent steps in the formation of this novel cyclopropyl amino acid.

Adenosine Monophosphate↗
PubMed2006

Type III effector diversification via both pathoadaptation and horizontal transfer in response to a coevolutionary arms race.

The concept of the coevolutionary arms race holds a central position in our understanding of pathogen-host interactions. Here we identify the molecular mechanisms and follow the stepwise progression of an arms race in a natural system. We show how the evolution and function of the HopZ family of type III secreted effector proteins carried by the plant pathogen Pseudomonas syringae are influenced by a coevolutionary arms race between pathogen and host. We surveyed 96 isolates of P. syringae and identified three homologs (HopZ1, HopZ2, and HopZ3) distributed among approximately 45% of the strains. All alleles were sequenced and their expression was confirmed. Evolutionary analyses determined that the diverse HopZ1 homologs are ancestral to P. syringae, and have diverged via pathoadaptive mutational changes into three functional and two degenerate forms, while HopZ2 and HopZ3 have been brought into P. syringae via horizontal transfer from other ecologically similar bacteria. A PAML selection analysis revealed that the C terminus of HopZ1 is under strong positive selection. Despite the extensive genetic variation observed in this family, all three homologs have cysteine-protease activity, although their substrate specificity may vary. The introduction of the ancestral hopZ1 allele into strains harboring alternate alleles results in a resistance protein-mediated defense response in their respective hosts, which is not observed with the endogenous allele. These data indicate that the P. syringae HopZ family has undergone allelic diversification via both pathoadaptive mutational changes and horizontal transfer in response to selection imposed by the host defense system. This genetic diversity permits the pathogen to avoid host defenses while still maintaining a virulence-associated protease, thereby allowing it to thrive on its current host, while simultaneously impacting its host range.

Adaptation, Biological↗