Isolation and characterization of adhesin-defective TnphoA mutants of septicaemic porcine Escherichia coli of serotype O115:K--: F165

Harel, Josée; Forget, Céline; Ngeleka, Musangu; Jacques, Mario; Fairbrother, John M.

doi:10.1099/00221287-138-11-2337

Cited by 16 publications

(20 citation statements)

References 36 publications

(29 reference statements)

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“…The Pho regulon has functions related to virulence in extraintestinal pathogenic E. coli and intestinal pathogenic E. coli. In the former, mutations in the phosphate-specific transport system led to reduced serum survival, reduced capsular antigen at the cell surface, changes in lipid A composition of the outer membrane, and an imbalance of cyclopropane and unsaturated fatty acids (59)(60)(61)(63)(64)(65). These changes contribute to virulence defects in a newborn pig infection model.…”

Section: Discussionmentioning

confidence: 99%

Defects in Phosphate Acquisition and Storage Influence Virulence of Cryptococcus neoformans

et al. 2014

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Nutrient acquisition and sensing are critical aspects of microbial pathogenesis. Previous transcriptional profiling indicated that the fungal pathogen Cryptococcus neoformans, which causes meningoencephalitis in immunocompromised individuals, encounters phosphate limitation during proliferation in phagocytic cells. We therefore tested the hypothesis that phosphate acquisition and polyphosphate metabolism are important for cryptococcal virulence. Deletion of the high-affinity uptake system interfered with growth on low-phosphate medium, perturbed the formation of virulence factors (capsule and melanin), reduced survival in macrophages, and attenuated virulence in a mouse model of cryptococcosis. Additionally, analysis of nutrient sensing functions for C. neoformans revealed regulatory connections between phosphate acquisition and storage and the iron regulator Cir1, cyclic AMP (cAMP)-dependent protein kinase A (PKA), and the calcium-calmodulin-activated protein phosphatase calcineurin. Deletion of the VTC4 gene encoding a polyphosphate polymerase blocked the ability of C. neoformans to produce polyphosphate. The vtc4 mutant behaved like the wild-type strain in interactions with macrophages and in the mouse infection model. However, the fungal load in the lungs was significantly increased in mice infected with vtc4 deletion mutants. In addition, the mutant was impaired in the ability to trigger blood coagulation in vitro, a trait associated with polyphosphate. Overall, this study reveals that phosphate uptake in C. neoformans is critical for virulence and that its regulation is integrated with key signaling pathways for nutrient sensing.

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Section: Discussionmentioning

confidence: 99%

Defects in Phosphate Acquisition and Storage Influence Virulence of Cryptococcus neoformans

et al. 2014

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“…Mutations were obtained from random insertion of the TnphoA sequence into the chromosomal DNA of E. coli strain 86-1390 (Sm r Tc r ). This was accomplished as described previously (17) by using the suicide vector pRT733, which carries the TnphoA insertion and the kanamycin resistance (Km r ) gene in E. coli strain S⌴10pir (51). Exconjugants from the mating between E. coli strain S⌴10pir(pRT733) and E. coli strain 86-1390 were selected on Luria-Bertani (LB) agar (Difco Laboratories, Detroit, Mich.) containing kanamycin and streptomycin (40 g ml Ϫ1 ) and the alkaline phosphatase substrate XP (5-bromo-4-chloro-3-indolylphosphate) (Sigma Chemical Co., St. Louis, Mo.).…”

Section: Methodsmentioning

confidence: 99%

“…A collection of 11 PEPEC strains was used for in vivo experiments. E. coli strain SM10pir(pRT733) was used to introduce TnphoA into strain 86-1390 by conjugation (17). E. coli strain HB101 (supE44 hsdS20 (r Ϫ m Ϫ ) recA13 ara-14 proA2 lacY1 galK2 rpsL20 xyl-5 mtl-1) (7) was used as host for recombinant plasmids in this study.…”

Section: Methodsmentioning

confidence: 99%

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Characterization of the Novel Factor Paa Involved in the Early Steps of the Adhesion Mechanism of Attaching and Effacing Escherichia coli

et al. 2003

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Nonenterotoxigenic porcine Escherichia coli strains belonging to the serogroup O45 have been associated with postweaning diarrhea in swine and adhere to intestinal epithelial cells in a characteristic attaching and effacing (A/E) pattern. O45 porcine enteropathogenic E. coli (PEPEC) strain 86-1390 induces typical A/E lesions in a pig ileal explant model. Using TnphoA transposon insertion mutagenesis on strain 86-1390, we found a mutant that did not induce A/E lesions. The insertion was identified in a gene designated paa (porcine A/E-associated gene). Sequence analysis of paa revealed an open reading frame of 753 bp encoding a 27.6-kDa protein which displayed 100, 51.8, and 49% homology with Paa of enterohemorrhagic E. coli O157:H7 strains (EDL933 and Sakai), PEB3 of Campylobacter jejuni, and AcfC of Vibrio cholerae, respectively. Chromosomal localization studies indicated that the region containing paa was inserted between the yciD and yciE genes at about 28.3 min of the E. coli K-12 chromosome. The presence of paa and eae sequences in the porcine O45 strains is highly correlated with the A/E phenotype. However, the observation that three eae-positive but paa-negative PEPEC O45 strains were A/E negative provides further evidence for the importance of the paa gene in the A/E activity of O45 strains. As well, the complementation of the paa mutant restored the A/E activity of the 86-1390 strain, showing the involvement of Paa in PEPEC pathogenicity. These observations suggest that Paa contributes to the early stages of A/E E. coli virulence.Attaching and effacing (A/E) Escherichia coli (AEEC) induces distinctive histopathological lesions on the intestinal mucosa, known as the A/E lesions. These lesions are characteristic of enteric pathogens such as enteropathogenic E. coli (EPEC), responsible for severe childhood diarrhea in developing countries (14, 38), enterohemorrhagic E. coli (EHEC), causing hemorrhagic colitis and hemolytic-uremic syndrome, a diarrheagenic E. coli strain of rabbits (RDEC-1), strains of Hafnia alvei isolated from children with diarrhea, and Citrobacter rodentium, causing transmissible colonic hyperplasia in mice (4,16,53). A/E lesions have also been associated with diarrhea in different animal species such as rabbits, calves, dogs, cats, lambs, pigs, and tamarins (8,9,22,32,37,55).A/E lesions result from intimate bacterial adherence to the apical surfaces of enterocytes and activation of several chromosomal gene products that interact with components of the host cell, leading to host cell protein phosphorylation, effacement of target brush borders, and disruption of the underlying actin cytoskeleton (11, 38). The genes are clustered in a chromosomal pathogenicity island called the locus of enterocyte effacement (LEE). Its location and size vary in different strains. In EPEC strain E2348/69 and EHEC O157:H7 strains, the LEE is inserted in the selC locus at about 82 min on the E. coli K-12 chromosome, but its size varies from 35 kb for EPEC to 43 kb for EHEC. In strains of serotype O26:H-, the...

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“…Grids were washed and placed on drops of anti-rabbit immunoglobulin with 10 nm gold particles (Sigma) for 30 min. After a final washing step, the grids were stained with 1 % (w/v) phosphotungstate (pH 7.0) and examined with an electron microscope (Philips 201) at an accelerating voltage of 60 kV (Harel et al, 1992a E. coli F165, fimbriae plates as described previously (Harel et al, 1992b). After boiling the fimbrial samples for 5 min in buffer [lo mM Tris/HCl (pH 7*8)] containing 4% (w/v) SDS, 0.01 ml p-mercaptoethanol, 0.2 ml glycerol and 0.002 % bromophenol blue, the samples were run on slab gels as described previously (Harel e t al., 1992a).…”

Section: Methodsmentioning

confidence: 99%

Cloning of determinants encoding F1652 fimbriae from porcine septicaemic Escherichia coli confirms their identity as F1C fimbriae

Harel

Jacques²,

Fairbrother³

et al. 1995

Microbiology

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Foy, Quebec, Canada GlV4G5 ~ Cloning of the f165, operon that encodes F1C-like fimbriae in Escherichia coli indicates that this operon is a member of the S/Foc family. The genetic determinant coding for the F165, fimbriae was cloned from the chromosome of the porcine E. coli wild-type strain 4787 (01 15: K-l : H51: F165). The cloned F165, and the wild-type operon expressed a major fimbrial protein subunit of molecular mass 17.2 kDa that was detected by anti-F165 and anti-FlC polyclonal sera. The sequences of the f165A and f165JGH genes are reported. Major subunit gene f165# encodes a mature protein of 156 amino acids. Minor subunit genes f, G and H encode mature proteins of 148,145 and 276 amino acids, respectively. The amino acid sequences of the four proteins share similarities with those of the known S and F1C fimbrial antigens that are produced by extraintestinal E. coli which is associated with sepsis, urinary tract infections and newborn meningitis. The F165,A protein was identical to the major subunit protein of FlC, with a difference only at the first position. It was also similar, to a lesser extent, to the major subunit proteins of Sfal and Sfall fimbriae. F165,F was identical to FocF and SfaWSfallG. F165,G was more closely related to FocG than to SfalS/SfallS, and F165,H was more closely related to FocH than to SfalH/SfallH.

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Isolation and characterization of adhesin-defective TnphoA mutants of septicaemic porcine Escherichia coli of serotype O115:K--: F165

Abstract: Non-enterotoxigenic porcine

Cited by 16 publications

References 36 publications

Defects in Phosphate Acquisition and Storage Influence Virulence of Cryptococcus neoformans

Defects in Phosphate Acquisition and Storage Influence Virulence of Cryptococcus neoformans

Characterization of the Novel Factor Paa Involved in the Early Steps of the Adhesion Mechanism of Attaching and Effacing Escherichia coli

Cloning of determinants encoding F1652 fimbriae from porcine septicaemic Escherichia coli confirms their identity as F1C fimbriae

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