scholarly journals Bradyrhizobium elkanii rtxC Gene Is Required for Expression of Symbiotic Phenotypes in the Final Step of Rhizobitoxine Biosynthesis

2004 ◽  
Vol 70 (1) ◽  
pp. 535-541 ◽  
Author(s):  
Shin Okazaki ◽  
Masayuki Sugawara ◽  
Kiwamu Minamisawa
Keyword(s):  
Glycine Max ◽  
Type Strain ◽  
Wild Type Strain ◽  
Null Mutant ◽  
Wild Type ◽  
In Planta ◽  
Distinct Effect ◽  
Nodulation Competitiveness ◽  
Macroptilium Atropurpureum ◽  
Bradyrhizobium Elkanii

ABSTRACT We disrupted the rtxC gene on the chromosome of Bradyrhizobium elkanii USDA94 by insertion of a nonpolar aph cartridge. The rtxC mutant, designated ΔrtxC, produced serinol and dihydrorhizobitoxine but no rhizobitoxine, both in culture and in planta. The introduction of cosmids harboring the rtxC gene into the ΔrtxC mutant complemented rhizobitoxine production, suggesting that rtxC is involved in the final step of rhizobitoxine biosynthesis in B. elkanii USDA94. Glycine max cv. Lee inoculated with ΔrtxC or with a null mutant, Δrtx::Ω1, showed no foliar chlorosis, whereas the wild-type strain USDA94 caused severe foliar chlorosis. The two mutants showed significantly less nodulation competitiveness than the wild-type strain on Macroptilium atropurpureum. These results indicate that dihydrorhizobitoxine, the immediate precursor of the oxidative form of rhizobitoxine, has no distinct effect on nodulation phenotype in these legumes. Thus, desaturation of dihydrorhizobitoxine by rtxC-encoded protein is essential for the bacterium to show rhizobitoxine phenotypes in planta. In addition, complementation analysis of rtxC by cosmids differing in rtxC transcription levels suggested that rhizobitoxine production correlates with the amount of rtxC transcript.

scholarly journals Expression of Erwinia chrysanthemi Pectinase Genes pelI, pelL, and pelZ During Infection of Potato Tubers

1999 ◽  
Vol 12 (10) ◽  
pp. 845-851 ◽  
Author(s):  
Sylwia Jafra ◽  
Izabela Figura ◽  
Nicole Hugouvieux-Cotte-Pattat ◽  
Ewa Lojkowska
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Pectate Lyase ◽  
Soft Rot ◽  
Erwinia Chrysanthemi ◽  
Potato Tubers ◽  
Wild Type ◽  
In Planta ◽  
Gene Encoding ◽  
Glucuronidase Activity

Erwinia chrysanthemi mutants, containing transcriptional fusions of one of the minor pectate lyase genes (pelI, pelL, pelZ) with the reporter gene encoding β-glucuronidase activity, were studied for their ability to cause disease symptoms and to synthesize pectinases after inoculation of potato tubers. The strains affected in pelI and pelL genes displayed reduced virulence on potato tubers, demonstrating the important role of these isoenzymes in soft rot disease. Inactivation of the pelZ gene slightly influences the ability to macerate. Analysis of the bacterial population showed rapid multiplication of bacteria during infection. Similar kinetics of growth were observed for all mutants and for the wild-type strain. Comparison of the mutants and the wild-type strain showed that the pelI, pelL, and pelZ mutants synthesized reduced levels of Pels. The expression of pelZ is fivefold higher in planta than in bacterial cultures. In contrast, both pelI and pelL are highly (10-fold factor) induced in planta, which is characteristic of the plant-inducible pectate lyases.

scholarly journals Quorum Sensing Regulates Exopolysaccharide Production, Motility, and Virulence in Pseudomonas syringae

2005 ◽  
Vol 18 (7) ◽  
pp. 682-693 ◽  
Author(s):  
Beatriz Quiñones ◽  
Glenn Dulla ◽  
Steven E. Lindow
Keyword(s):  
Quorum Sensing ◽  
Pseudomonas Syringae ◽  
Type Strain ◽  
Double Mutant ◽  
Wild Type Strain ◽  
Homoserine Lactone ◽  
Brown Spot ◽  
Wild Type ◽  
In Planta ◽  
Alginate Production

The N-acyl homoserine lactone (AHL)-mediated quorumsensing system in the phytopathogen Pseudomonas syringae pv. syringae requires the AHL synthase AhlI and the regulator AhlR, and is additionally subject to regulation by AefR. The contribution of quorum sensing to the expression of a variety of traits expected to be involved in epiphytic fitness and virulence of P. syringae were examined. Both an aefR- mutant and an ahlR- double mutant, deficient in AHL production, were significantly impaired in alginate production and had an increased susceptibility to hydrogen peroxide compared with the wild-type strain. These mutants were hypermotile in culture, invaded leaves more rapidly, and caused an increased incidence of brown spot lesions on bean leaves after a 48-h moist incubation. Interestingly, an aefR- mutant was both the most motile and virulent. Like the wild-type strain, the AHL-deficient mutant strains incited water-soaked lesions on bean pods. However, lesions caused by an ahlI- ahlR- double mutant were larger, whereas those incited by an aefR- mutant were smaller. In contrast, tissue maceration of pods, which occurs at a later stage of infection, was completely abolished in the AHL-deficient mutants. Both the incidence of disease and in planta growth of P. syringae pv. tabaci were greatly reduced in transgenic tobacco plants that produced AHL compared with wild-type plants. These results demonstrate that quorum sensing in P. syringae regulates traits that contribute to epiphytic fitness as well as to distinct stages of disease development during plant infection.

scholarly journals The Role of Ethylene Production in Virulence of Pseudomonas syringae pvs. glycinea and phaseolicola

2001 ◽  
Vol 91 (5) ◽  
pp. 511-518 ◽  
Author(s):  
Helge Weingart ◽  
Henriette Ullrich ◽  
Klaus Geider ◽  
Beate Völksch
Keyword(s):  
Pseudomonas Syringae ◽  
Ethylene Production ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
In Planta ◽  
Population Sizes ◽  
Soybean Plants ◽  
Type Strains

The importance of ethylene production for virulence of Pseudomonas syringae pvs. glycinea and phaseolicola was assayed by comparing bacterial multiplication and symptom development in bean and soybean plants inoculated with ethylene-negative (efe) mutants and wild-type strains. The efe mutants of Pseudomonas syringae pv. glycinea were significantly reduced in their ability to grow in planta. However, the degree of reduction was strain-dependent. Population sizes of efe mutant 16/83-E1 that did not produce the phototoxin coronatine were 10- and 15-fold lower than those of the wild-type strain on soybean and on bean, and 16/83-E1 produced very weak symptoms compared with the wild-type strain. The coronatine-producing efe mutant 7a/90-E1 reached fourfold and twofold lower population sizes compared with the wild-type strain on soybean and bean, respectively, and caused disease symptoms typical of the wild-type strain. Experiments with ethylene-insensitive soybeans confirmed these results. The virulence of the wild-type strains was reduced to the same extent in ethylene-insensitive soybean plants as the virulence of the efe mutants in ethylene-susceptible soybeans. In contrast, the virulence of Pseudomonas syringae pv. phaseolicola was not affected by disruption of the efe gene.

scholarly journals Mutants Suppressing Novobiocin Hypersensitivity of a mukB Null Mutation

2003 ◽  
Vol 185 (13) ◽  
pp. 3690-3695 ◽  
Author(s):  
Shun Adachi ◽  
Sota Hiraga
Keyword(s):  
Chromosome Segregation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Dna Gyrase ◽  
Beta Subunit ◽  
Null Mutation ◽  
Null Mutant ◽  
Wild Type ◽  
Suppressor Mutations ◽  
Sister Chromosome

ABSTRACT The mukB gene is essential for the partitioning of sister chromosomes in Escherichia coli. A mukB null mutant is hypersensitive to the DNA gyrase inhibitor novobiocin. In this work, we isolated mutants suppressing the novobiocin hypersensitivity of the mukB null mutation. All suppressor mutations are localized in or near the gyrB gene, and the four tested clones have an amino acid substitution in the DNA gyrase beta subunit. We found that in the mukB mutant, the process of sister chromosome segregation is strikingly hypersensitive to novobiocin; however, the effect of novobiocin on growth, which was measured by culture turbidity, is the same as that of the wild-type strain.

scholarly journals The In Planta-Produced Extracellular Proteins ECP1 and ECP2 of Cladosporium fulvum Are Virulence Factors

1997 ◽  
Vol 10 (6) ◽  
pp. 725-734 ◽  
Author(s):  
Richard Laugé ◽  
Matthieu H. A. J. Joosten ◽  
Guido F. J. M. Van den Ackerveken ◽  
Henk W. J. Van den Broek ◽  
Pierre J. G. M. De Wit
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Pathogenic Fungus ◽  
Leaf Tissue ◽  
Defense Responses ◽  
Gene Replacement ◽  
Extracellular Proteins ◽  
Cladosporium Fulvum ◽  
Wild Type ◽  
In Planta

The two extracellular proteins ECP1 and ECP2 are abundantly secreted by the plant-pathogenic fungus Cladosporium fulvum during colonization of the intercellular space of tomato leaves. We examined the involvement of both proteins in pathogenicity and virulence of this fungus. ECP1-deficient, ECP2-deficient, and ECP1/ECP2- deficient isogenic C. fulvum strains were created by targeted gene replacement. Upon inoculation onto susceptible 6-week-old tomato plants, all three mutants showed reduced virulence. Deficiency in ECP2 resulted in a strain that poorly colonized the leaf tissue and secreted lower amounts of the in planta-produced ECP3, AVR4, and AVR9 proteins than the wild-type strain. The ECP2-deficient strain produced little emerging mycelium and few conidia. Deficiency in ECP1 did not significantly modify colonization of the leaf tissue, but reduced secretion of in planta-produced proteins. The ECP1-deficient strain emerged from stomata of the lower epidermis, but failed to sporulate as abundantly as the wild-type strain. A strain deficient in both ECP1 and ECP2 proteins had a phenotype similar to that of the ECP2-deficient strain. Accumulation of pathogenesis-related proteins and induction of late responses, such as leaf desiccation and abscission, occurred more quickly and more severely in tomato after inoculation with the ECP1-, ECP2-, and ECP1/ECP2-deficient strains than after inoculation with the wild-type strain. Moreover, partial collapse of stomatal guard cells occurred at emergence of the ECP2-deficient strain. These results indicate that the ECP1 and ECP2 proteins play a role in virulence of C. fulvum on tomato and suggest that both are involved in suppression of host defense responses.

scholarly journals Hypovirulence-associated traits induced by a mycovirus of Cryphonectria parasitica are mimicked by targeted inactivation of a host gene.

1993 ◽  
Vol 13 (12) ◽  
pp. 7782-7792 ◽  
Author(s):  
L Zhang ◽  
A C Churchill ◽  
P Kazmierczak ◽  
D H Kim ◽  
N K Van Alfen
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Cryphonectria Parasitica ◽  
Apple Fruit ◽  
Host Gene ◽  
Fruiting Bodies ◽  
Null Mutant ◽  
Wild Type ◽  
Fungal Virulence ◽  
Double Stranded Rna

Expression of the Vir2 gene of Cryphonectria parasitica is down-regulated in strains of the fungus containing a double-stranded RNA genetic element that reduces fungal virulence (W. A. Powell and N. K. Van Alfen, Mol. Cell. Biol. 7:3688-3693, 1987). We have sequenced the Vir2 gene and characterized its structure; the mRNA contains a short open reading frame whose product has structural similarities to several fungal pheromones. A null mutant was constructed by homologous recombination to determine the function of the Vir2 gene and whether its disruption resulted in any of the altered phenotypes exhibited by many hypovirulent strains, such as reductions in virulence, pigmentation, and sporulation. The Vir2 null mutant (18dm) exhibited a wild-type phenotype with respect to gross colony morphology, growth rate, pigmentation, asexual spore viability, and virulence in apple fruit and chestnut trees. However, numbers of asexual fruiting bodies (pycnidia) and conidia were reduced significantly in comparison with the wild-type strain EP155/2. In sexual crosses of 18dm with a wild-type strain of the opposite mating type, perithecia (sexual fruiting bodies) developed but were barren. Deletion of the Vir2 gene results in a phenotype that mimics that of many double-stranded-RNA-containing hypovirulent strains; i.e., the null mutant exhibits significant reductions in asexual sporulation and pycinidum production as well as impaired sexual crossing ability. To our knowledge, this is the first report of the partial reproduction of a virus-induced phenotype by deletion of a virus-perturbed host gene.

scholarly journals The CreC Regulator of Escherichia coli, a New Target for Metabolic Manipulations

2015 ◽  
Vol 82 (1) ◽  
pp. 244-254 ◽  
Author(s):  
Manuel S. Godoy ◽  
Pablo I. Nikel ◽  
José G. Cabrera Gomez ◽  
M. Julia Pettinari
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Wild Type Strain ◽  
Oxygen Availability ◽  
Biochemical Network ◽  
Metabolic Effects ◽  
Null Mutant ◽  
Wild Type ◽  
Content Type ◽  
E Coli

ABSTRACTThe CreBC (carbon source-responsive) two-component regulation system ofEscherichia coliaffects a number of functions, including intermediary carbon catabolism. The impacts of differentcreCmutations (a ΔcreCmutant and a mutant carrying the constitutivecreC510allele) on bacterial physiology were analyzed in glucose cultures under three oxygen availability conditions. Differences in the amounts of extracellular metabolites produced were observed in the null mutant compared to the wild-type strain and the mutant carryingcreC510and shown to be affected by oxygen availability. The ΔcreCstrain secreted more formate, succinate, and acetate but less lactate under low aeration. These metabolic changes were associated with differences in AckA and LdhA activities, both of which were affected by CreC. Measurement of the NAD(P)H/NAD(P)+ratios showed that thecreC510strain had a more reduced intracellular redox state, while the opposite was observed for the ΔcreCmutant, particularly under intermediate oxygen availability conditions, indicating that CreC affects redox balance. The null mutant formed more succinate than the wild-type strain under both low aeration and no aeration. Overexpression of the genes encoding phosphoenolpyruvate carboxylase fromE. coliand a NADH-forming formate dehydrogenase fromCandida boidiniiin the ΔcreCmutant further increased the yield of succinate on glucose. Interestingly, the elimination ofackAandadhEdid not significantly improve the production of succinate. The diverse metabolic effects of this regulator on the central biochemical network ofE. colimake it a good candidate for metabolic-engineering manipulations to enhance the formation of bioproducts, such as succinate.

scholarly journals Epsilon Toxin Is Essential for the Virulence of Clostridium perfringens Type D Infection in Sheep, Goats, and Mice

2013 ◽  
Vol 81 (7) ◽  
pp. 2405-2414 ◽  
Author(s):  
J. P. Garcia ◽  
V. Adams ◽  
J. Beingesser ◽  
M. L. Hughes ◽  
R. Poon ◽  
...  
Keyword(s):  
Clostridium Perfringens ◽  
Type Strain ◽  
Wild Type Strain ◽  
Null Mutant ◽  
Wild Type ◽  
Histological Changes ◽  
Content Type ◽  
Type D ◽  
Necrotizing Colitis ◽  
Epsilon Toxin

ABSTRACTClostridium perfringenstype D causes disease in sheep, goats, and other ruminants. Type D isolates produce, at minimum, alpha and epsilon (ETX) toxins, but some express up to five different toxins, raising questions about which toxins are necessary for the virulence of these bacteria. We evaluated the contribution of ETX toC. perfringenstype D pathogenicity in an intraduodenal challenge model in sheep, goats, and mice using a virulentC. perfringenstype D wild-type strain (WT), an isogenic ETX null mutant (etxmutant), and a strain where theetxmutation has been reversed (etxcomplemented). All sheep and goats, and most mice, challenged with the WT isolate developed acute clinical disease followed by death in most cases. Sheep developed various gross and/or histological changes that included edema of brain, lungs, and heart as well as hydropericardium. Goats developed various effects, including necrotizing colitis, pulmonary edema, and hydropericardium. No significant gross or histological abnormalities were observed in any mice infected with the WT strain. All sheep, goats, and mice challenged with the isogenicetxmutant remained clinically healthy for ≥24 h, and no gross or histological abnormalities were observed in those animals. Complementation ofetxknockout restored virulence; most goats, sheep, and mice receiving this complemented mutant developed clinical and pathological changes similar to those observed in WT-infected animals. These results indicate that ETX is necessary for type D isolates to induce disease, supporting a key role for this toxin in type D disease pathogenesis.

scholarly journals Molecular Monitoring of Wild-Type and Genetically Engineered Colletotrichum coccodes Biocontrol Strains In Planta

Plant Disease ◽  
2006 ◽  
Vol 90 (12) ◽  
pp. 1504-1510 ◽  
Author(s):  
A. L. Dauch ◽  
B. Ahn ◽  
A. K. Watson ◽  
P. Seguin ◽  
S. H. Jabaji-Hare
Keyword(s):  
Type Strain ◽  
Wild Type Strain ◽  
Quantitative Polymerase Chain Reaction ◽  
Fungal Growth ◽  
Genetically Engineered ◽  
Colletotrichum Coccodes ◽  
Wild Type ◽  
In Planta ◽  
Molecular Monitoring ◽  
Dna Amounts

Two strains of Colletotrichum coccodes, the wild type (DAOM 183088) and T-20a, engineered with the necrosis- and ethylene-inducing peptide (NEP1) gene for hypervirulence on velvetleaf (Abutilon theophrasti, Medik.), were monitored in planta for the first 2 weeks after infection. Real-time quantitative polymerase chain reaction (QPCR) was used to assess the extent of colonization of both strains on velvetleaf using SYBR Green chemistry. Quantification of both strains was successful as soon as the conidia were sprayed on the leaves and up to 14 days after infection. The increase in fungal DNA amounts corroborated with the appearance of necrotic lesions on velvetleaf leaves infected with the wild-type strain. The wild-type C. coccodes was more efficient at infecting velvetleaf than the transgenic T-20a strain. In addition, detection of host DNA allowed us to quantitatively monitor the decrease in plant DNA amounts in response to wild-type strain infection. Expression of the NEP1 transgene by conventional retro-transcription (RT)-PCR was absent from T-20a growing on either V8 agar or in planta, suggesting that the gene may be silenced. The application of QPCR to monitor fungal growth was proven to detect the target organisms in planta prior to the appearance of symptoms.

scholarly journals Requirement of the galU Gene for Polysaccharide Production by and Pathogenicity and Growth In Planta of Xanthomonas citri subsp. citri

2010 ◽  
Vol 76 (7) ◽  
pp. 2234-2242 ◽  
Author(s):  
Yinping Guo ◽  
Uma Shankar Sagaram ◽  
Jeong-soon Kim ◽  
Nian Wang
Keyword(s):  
Biofilm Formation ◽  
Type Strain ◽  
Virulence Genes ◽  
Wild Type Strain ◽  
Capsular Polysaccharide ◽  
Extracellular Polysaccharides ◽  
Wild Type ◽  
In Planta ◽  
Intercellular Spaces ◽  
Xanthomonas Citri

ABSTRACT Xanthomonas citri subsp. citri is the causal agent of citrus canker, which has a significant impact on citrus production. In this study, we characterized the galU gene of X. citri subsp. citri. Two galU mutants (F6 and D12) were identified in an X. citri subsp. citri EZ-Tn5 <R6Kγori/KAN-2> Tnp transposon library. Rescue cloning, sequence analysis, and Southern blot analysis indicated that both of these mutants had a single copy of the EZ-Tn5 transposon inserted in galU in the chromosome. Further study showed that galU was required for biosynthesis of extracellular polysaccharides (EPS; xanthan gum) and capsular polysaccharide (CPS) and biofilm formation. Mutation of galU resulted in a loss of pathogenicity for grapefruit. The loss of pathogenicity of a galU mutant resulted from its inability to grow in planta rather than from the effect on virulence genes. Quantitative reverse transcription-PCR assays indicated that mutation of galU did not impair the expression of key virulence genes, such as pthA of X. citri subsp. citri. Although D12 had a growth rate similar to that of the wild-type strain in nutrient broth, no D12 population became established in the intercellular spaces of citrus leaves. Coinoculation of a galU mutant with the wild-type strain did not promote growth of the galU mutant in planta. Defects in EPS and CPS production, pathogenicity, and growth in planta of the galU mutant were complemented to the wild-type level using plasmid pCGU2.1 containing an intact galU gene. These data indicate that the galU gene contributes to X. citri subsp. citri growth in intercellular spaces and is involved in EPS and CPS synthesis and biofilm formation.

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