scholarly journals The Escherichia coli Fis Protein Stimulates Bacteriophage λ Integrative Recombination In Vitro

2003 ◽  
Vol 185 (10) ◽  
pp. 3076-3080 ◽  
Author(s):  
Dominic Esposito ◽  
Gary F. Gerard
Keyword(s):  
Escherichia Coli ◽  
Host Cell ◽  
Essential Role ◽  
Bacteriophage Λ ◽  
Site Specific ◽  
Site Specific Recombination ◽  
E Coli ◽  
In Vivo Recombination

ABSTRACT The Escherichia coli nucleoid-associated protein Fis was previously shown to be involved in bacteriophage lambda site-specific recombination in vivo, enhancing the levels of both integrative recombination and excisive recombination. While purified Fis protein was shown to stimulate in vitro excision, Fis appeared to have no effect on in vitro integration reactions even though a 15-fold drop in lysogenization frequency had previously been observed in fis mutants. We demonstrate here that E. coli Fis protein does stimulate integrative lambda recombination in vitro but only under specific conditions which likely mimic natural in vivo recombination more closely than the standard conditions used in vitro. In the presence of suboptimal concentrations of Int protein, Fis stimulates the rate of integrative recombination significantly. In addition, Fis enhances the recombination of substrates with nonstandard topologies which may be more relevant to the process of in vivo phage lambda recombination. These data support the hypothesis that Fis may play an essential role in lambda recombination in the host cell.

scholarly journals Mutual Enhancement of Virulence by Enterotoxigenic and Enteropathogenic Escherichia coli

2006 ◽  
Vol 74 (3) ◽  
pp. 1505-1515 ◽  
Author(s):  
John K. Crane ◽  
Shilpa S. Choudhari ◽  
Tonniele M. Naeher ◽  
Michael E. Duffey
Keyword(s):  
Escherichia Coli ◽  
Host Cell ◽  
Synergistic Effects ◽  
Adenine Nucleotides ◽  
Ussing Chamber ◽  
Atp Release ◽  
T84 Cells ◽  
E Coli

ABSTRACT Enterotoxigenic Escherichia coli (ETEC) and enteropathogenic E. coli (EPEC) are common causes of diarrhea in children in developing countries. Dual infections with both pathogens have been noted fairly frequently in studies of diarrhea around the world. In previous laboratory work, we noted that cholera toxin and forskolin markedly potentiated EPEC-induced ATP release from the host cell, and this potentiated release was found to be mediated by the cystic fibrosis transmembrane conductance regulator. In this study, we examined whether the ETEC heat-labile toxin (LT) or the heat-stable toxin (STa, also known as ST) potentiated EPEC-induced ATP release. We found that crude ETEC culture filtrates, as well as purified ETEC toxins, did potentiate EPEC-induced ATP release in cultured T84 cells. Coinfection of T84 cells with live ETEC plus EPEC bacteria also resulted in enhanced ATP release compared to EPEC alone. In Ussing chamber studies of chloride secretion, adenine nucleotides released from the host by EPEC also significantly enhanced the chloride secretory responses that were triggered by crude ETEC filtrates, purified STa, and the peptide hormone guanylin. In addition, adenosine and LT had additive or synergistic effects in inducing vacuole formation in T84 cells. Therefore, ETEC toxins and EPEC-induced damage to the host cell both enhance the virulence of the other type of E. coli. Our in vitro data demonstrate a molecular basis for a microbial interaction, which could result in increased severity of disease in vivo in individuals who are coinfected with ETEC and EPEC.

scholarly journals The ripX Locus of Bacillus subtilis Encodes a Site-Specific Recombinase Involved in Proper Chromosome Partitioning

1999 ◽  
Vol 181 (19) ◽  
pp. 6053-6062 ◽  
Author(s):  
Stephen A. Sciochetti ◽  
Patrick J. Piggot ◽  
David J. Sherratt ◽  
Garry Blakely
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Holliday Junction ◽  
Independent Manner ◽  
Site Specific ◽  
E Coli ◽  
Chromosome Partitioning ◽  
A Site

ABSTRACT The Bacillus subtilis ripX gene encodes a protein that has 37 and 44% identity with the XerC and XerD site-specific recombinases of Escherichia coli. XerC and XerD are hypothesized to act in concert at the dif site to resolve dimeric chromosomes formed by recombination during replication. Cultures of ripX mutants contained a subpopulation of unequal-size cells held together in long chains. The chains included anucleate cells and cells with aberrantly dense or diffuse nucleoids, indicating a chromosome partitioning failure. This result is consistent with RipX having a role in the resolution of chromosome dimers inB. subtilis. Spores contain a single uninitiated chromosome, and analysis of germinated, outgrowing spores showed that the placement of FtsZ rings and septa is affected in ripXstrains by the first division after the initiation of germination. The introduction of a recA mutation into ripXstrains resulted in only slight modifications of the ripXphenotype, suggesting that chromosome dimers can form in a RecA-independent manner in B. subtilis. In addition to RipX, the CodV protein of B. subtilis shows extensive similarity to XerC and XerD. The RipX and CodV proteins were shown to bind in vitro to DNA containing the E. coli dif site. Together they functioned efficiently in vitro to catalyze site-specific cleavage of an artificial Holliday junction containing adif site. Inactivation of codV alone did not cause a discernible change in phenotype, and it is speculated that RipX can substitute for CodV in vivo.

scholarly journals Lysyl-tRNA synthetase from Escherichia coli K12. Chromatographic heterogeneity and the lysU-gene product

1987 ◽  
Vol 248 (1) ◽  
pp. 43-51 ◽  
Author(s):  
J Charlier ◽  
R Sanchez
Keyword(s):  
Escherichia Coli ◽  
Gene Product ◽  
Activity Ratio ◽  
Deae Cellulose ◽  
Trna Synthetase ◽  
Trna Synthetases ◽  
E Coli ◽  
Aminoacyl Trna Synthetases

In contrast with most aminoacyl-tRNA synthetases, the lysyl-tRNA synthetase of Escherichia coli is coded for by two genes, the normal lysS gene and the inducible lysU gene. During its purification from E. coli K12, lysyl-tRNA synthetase was monitored by its aminoacylation and adenosine(5′)tetraphospho(5′)adenosine (Ap4A) synthesis activities. Ap4A synthesis was measured by a new assay using DEAE-cellulose filters. The heterogeneity of lysyl-tRNA synthetase (LysRS) was revealed on hydroxyapatite; we focused on the first peak, LysRS1, because of its higher Ap4A/lysyl-tRNA activity ratio at that stage. Additional differences between LysRS1 and LysRS2 (major peak on hydroxyapatite) were collected. LysRS1 was eluted from phosphocellulose in the presence of the substrates, whereas LysRS2 was not. Phosphocellulose chromatography was used to show the increase of LysRS1 in cells submitted to heat shock. Also, the Mg2+ optimum in the Ap4A-synthesis reaction is much higher for LysRS1. LysRS1 showed a higher thermostability, which was specifically enhanced by Zn2+. These results in vivo and in vitro strongly suggest that LysRS1 is the heat-inducible lysU-gene product.

scholarly journals Biological Cost of Single and Multiple Norfloxacin Resistance Mutations in Escherichia coli Implicated in Urinary Tract Infections

2005 ◽  
Vol 49 (6) ◽  
pp. 2343-2351 ◽  
Author(s):  
Patricia Komp Lindgren ◽  
Linda L. Marcusson ◽  
Dorthe Sandvang ◽  
Niels Frimodt-Møller ◽  
Diarmaid Hughes
Keyword(s):  
Escherichia Coli ◽  
Urinary Tract ◽  
Infection Model ◽  
Resistance Mutations ◽  
Topoisomerase Iv ◽  
E Coli ◽  
Tract Infections ◽  
Subsequent Selection

ABSTRACT Resistance to fluoroquinolones in urinary tract infection (UTIs) caused by Escherichia coli is associated with multiple mutations, typically those that alter DNA gyrase and DNA topoisomerase IV and those that regulate AcrAB-TolC-mediated efflux. We asked whether a fitness cost is associated with the accumulation of these multiple mutations. Mutants of the susceptible E. coli UTI isolate Nu14 were selected through three to five successive steps with norfloxacin. Each selection was performed with the MIC of the selected strain. After each selection the MIC was measured; and the regions of gyrA, gyrB, parC, and parE, previously associated with resistance mutations, and all of marOR and acrR were sequenced. The first selection step yielded mutations in gyrA, gyrB, and marOR. Subsequent selection steps yielded mutations in gyrA, parE, and marOR but not in gyrB, parC, or acrR. Resistance-associated mutations were identified in almost all isolates after selection steps 1 and 2 but in less than 50% of isolates after subsequent selection steps. Selected strains were competed in vitro, in urine, and in a mouse UTI infection model against the starting strain, Nu14. First-step mutations were not associated with significant fitness costs. However, the accumulation of three or more resistance-associated mutations was usually associated with a large reduction in biological fitness, both in vitro and in vivo. Interestingly, in some lineages a partial restoration of fitness was associated with the accumulation of additional mutations in late selection steps. We suggest that the relative biological costs of multiple mutations may influence the evolution of E. coli strains that develop resistance to fluoroquinolones.

scholarly journals The Escherichia coli cysG promoter belongs to the ‘extended −10’ class of bacterial promoters

1993 ◽  
Vol 296 (3) ◽  
pp. 851-857 ◽  
Author(s):  
T Belyaeva ◽  
L Griffiths ◽  
S Minchin ◽  
J Cole ◽  
S Busby
Keyword(s):  
Escherichia Coli ◽  
Point Mutations ◽  
Growth Conditions ◽  
Upstream Region ◽  
E Coli ◽  
Run Off ◽  
A Site ◽  
Specific Sequences

The Escherichia coli cysG promoter has been subcloned and shown to function constitutively in a range of different growth conditions. Point mutations identify the -10 hexamer and an important 5′-TGN-3′ motif immediately upstream. The effects of different deletions suggest that specific sequences in the -35 region are not essential for the activity of this promoter in vivo. This conclusion was confirmed by in vitro run-off transcription assays. The DNAase I footprint of RNA polymerase at the cysG promoter reveals extended protection upstream of the transcript start, and studies with potassium permanganate as a probe suggest that the upstream region is distorted in open complexes. Taken together, the results show that the cysG promoter belongs to the ‘extended -10’ class of promoters, and the base sequence is similar to that of the P1 promoter of the E. coli galactose operon, another promoter in this class. In vivo, messenger initiated at the cysG promoter appears to be processed by cleavage at a site 41 bases downstream from the transcript start point.

scholarly journals Evaluation du potentiel antimicrobien et de la toxicité des extraits de Jatropha multifida Linn, (Euphorbiaceae)

2020 ◽  
Vol 151 ◽  
pp. 15550-15558
Author(s):  
Amégninou Agban ◽  
Yao Hoekou ◽  
Passimna Pissang ◽  
Tchadjobo Tchacondo ◽  
Komlan Batawila
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Antimicrobial Activity ◽  
Candida Albicans ◽  
Aqueous Extract ◽  
E Coli ◽  
Toxicological Studies ◽  
Jatropha Multifida

Objectif : L’objectif de ce travail était d’évaluer in vitro l’activité antimicrobienne des extraits de feuilles et tige de Jatropha multifida sur la croissance de Candida albicans, Escherichia coli et Staphylococcus aureus, puis d’évaluer in vivo la toxicité de cette plante. Méthodologie et résultats : Les méthodes de diffusion en milieu gélosé et de microdilution en milieu liquide ont été utilisées pour évaluer l’effet antimicrobien. Une étude en subaigüe était réalisée afin d’explorer les effets toxiques de l’extrait aqueux des feuilles. Les résultats des tests antimicrobiens montrent une activité des extraits de feuilles et tige de J. multifida sur la croissance des souches utilisées avec des diamètres de zones d’inhibition allant de 8 à 25 mm et des concentrations minimales inhibitrices (CMI) variant de 0,039 mg/mL à 1,25 mg/mL à l’exception des souches de E. coli qui sont résistantes aux extraits de la tige. L’administration en subaigüe de l’extrait aqueux des feuilles de J. multifida à la dose de 600 mg/kg entraîne une perte significative de poids chez les souris. Conclusion et applications des résultats : Les extraits aqueux, éthanolique et hydroéthanolique des feuilles et tige de J. multifida possèdent d’activité antimicrobienne et pourraient être utilisés dans le traitement des Candidoses à C. albicans et des infections à S. aureus. Mais l’essai de toxicité subaigüe montre que l’extrait aqueux de la plante serait toxique. Des études toxicologiques approfondies restent donc nécessaires sur ces extraits afin de mieux élucider leur inocuité. Mots-clés : Jatropha multifida, extraits de feuilles et de tige, activités antifongique et antibactérienne, toxicité. Agban et al., J. Appl. Biosci. 2020 Evaluation du potentiel antimicrobien et de la toxicité des extraits de Jatropha multifida Linn, (Euphorbiaceae) 15551 Evaluation of antimicrobial potential and toxicity of Jatropha multifida Linn, (Euphorbiaceae) extracts ABSTRACT Objective: The objective of this study was to evaluate in vitro the antimicrobial activity of leaves and stem of Jatropha multifida extracts against Candida albicans, Escherichia coli and Staphylococcus aureus, and then to evaluate in vivo the toxicity of this plant. Methodology and Results: The agar well-diffusion and the NCCLS broth microdilution methods were used to assess the antimicrobial effect. A subacute study was carried out to explore the toxic effects of the aqueous extract of the leaves. The results of the antimicrobial tests show an activity of the extracts of leaves and stems of J. multifida on the growth of the strains used with diameters of inhibitory zones ranging from 8 to 25 mm and minimum inhibitory concentrations (MIC) varying from 0.039 mg/mL to 1.25 mg/mL exception E. coli strains which are resistant to extracts from the stem. Subacute administration of the aqueous extract of the leaves of J. multifida at a dose of 600 mg/kg leads to a significant loss of weight in the mice. Conclusion and application of findings : The aqueous, ethanolic and hydroethanolic extracts of the leaves and stem of J. multifida have antimicrobial activity and could be used in the treatment of Candidiasis and bacterial infections due respectively to C. albicans and S. aureus. But the subacute toxicity test shows that the aqueous extract of the plant would be toxic. Extensive toxicological studies therefore remain necessary on these extracts in order to better elucidate their safety. Keywords: Jatropha multifida extracts of leaves and stem, antifungal and antibacterial activities, toxicity

scholarly journals In Vitro and In Vivo Assessment of the Potential of Escherichia coli Phages to Treat Infections and Survive Gastric Conditions

2021 ◽  
Vol 9 (9) ◽  
pp. 1869
Author(s):  
Joanna Kaczorowska ◽  
Eoghan Casey ◽  
Gabriele A. Lugli ◽  
Marco Ventura ◽  
David J. Clarke ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Galleria Mellonella ◽  
Enterotoxigenic Escherichia Coli ◽  
E Coli ◽  
Composite Mixture ◽  
Ph Conditions ◽  
The Stability ◽  
Higher Sensitivity

Enterotoxigenic Escherichia coli (ETEC) and Shigella ssp. infections are associated with high rates of mortality, especially in infants in developing countries. Due to increasing levels of global antibiotic resistance exhibited by many pathogenic organisms, alternative strategies to combat such infections are urgently required. In this study, we evaluated the stability of five coliphages (four Myoviridae and one Siphoviridae phage) over a range of pH conditions and in simulated gastric conditions. The Myoviridae phages were stable across the range of pH 2 to 7, while the Siphoviridae phage, JK16, exhibited higher sensitivity to low pH. A composite mixture of these five phages was tested in vivo in a Galleria mellonella model. The obtained data clearly shows potential in treating E. coli infections prophylactically.

Effect of Tannins on the In Vitro Growth of Escherichia coli O157:H7 and In Vivo Growth of Generic Escherichia coli Excreted from Steers

2007 ◽  
Vol 70 (3) ◽  
pp. 543-550 ◽  
Author(s):  
BYENG R. MIN ◽  
WILLIAM E. PINCHAK ◽  
ROBIN C. ANDERSON ◽  
TODD R. CALLAWAY
Keyword(s):  
Escherichia Coli ◽  
Growth Rate ◽  
Bacterial Growth ◽  
Escherichia Coli O157 ◽  
Tannin Extract ◽  
Bacterial Growth Rate ◽  
Linear Effect ◽  
E Coli

The effect of commercially available chestnut and mimosa tannins in vitro (experiment 1) or in vivo (experiment 2) on the growth or recovery of Escherichia coli O157:H7 or generic fecal E. coli was evaluated. In experiment 1, the mean growth rate of E. coli O157:H7, determined via the measurement of optical density at 600 nm during anaerobic culture in tryptic soy broth at 37°C, was reduced (P < 0.05) with as little as 400 μg of either tannin extract per ml of culture fluid. The addition of 200, 400, 600, 800, and 1,200 μg of tannins per ml significantly (P < 0.01) reduced the specific bacterial growth rate when compared with the nontannin control. The specific growth rate decreased with increasing dose levels up to 800 μg of tannins per ml. Bacterial growth inhibition effects in chestnut tannins were less pronounced than in mimosa tannins. Chestnut tannin extract addition ranged from 0 to 1,200 μg/ml, and a linear effect (P < 0.05) was observed in cultures incubated for 6 h against the recovery of viable cells, determined via the plating of each strain onto MacConkey agar, of E. coli O157:H7 strains 933 and 86-24, but not against strain 6058. Similar tests with mimosa tannin extract showed a linear effect (P < 0.05) against the recovery of E. coli O157:H7 strain 933 only. The bactericidal effect observed in cultures incubated for 24 h with the tannin preparations was similar, although it was less than that observed from cultures incubated for 6 h. When chestnut tannins (15 g of tannins per day) were infused intraruminally to steers fed a Bermuda grass hay diet in experiment 2, fecal E. coli shedding was lower on days 3 (P < 0.03), 12 (P = 0.08), and 15 (P < 0.001) when compared with animals that were fed a similar diet without tannin supplementation. It was concluded that dietary levels and sources of tannins potentially reduce the shedding of E. coli from the gastrointestinal tract.

scholarly journals A Toxic Environment: a Growing Understanding of How Microbial Communities Affect Escherichia coli O157:H7 Shiga Toxin Expression

2020 ◽  
Vol 86 (24) ◽  
Author(s):  
Erin M. Nawrocki ◽  
Hillary M. Mosso ◽  
Edward G. Dudley
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Microbial Interactions ◽  
Severe Illness ◽  
Content Type ◽  
E Coli ◽  
Wide Range ◽  
Lambdoid Prophage

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) strains, including E. coli O157:H7, cause severe illness in humans due to the production of Shiga toxin (Stx) and other virulence factors. Because Stx is coregulated with lambdoid prophage induction, its expression is especially susceptible to environmental cues. Infections with Stx-producing E. coli can be difficult to model due to the wide range of disease outcomes: some infections are relatively mild, while others have serious complications. Probiotic organisms, members of the gut microbiome, and organic acids can depress Stx production, in many cases by inhibiting the growth of EHEC strains. On the other hand, the factors currently known to amplify Stx act via their effect on the stx-converting phage. Here, we characterize two interactive mechanisms that increase Stx production by O157:H7 strains: first, direct interactions with phage-susceptible E. coli, and second, indirect amplification by secreted factors. Infection of susceptible strains by the stx-converting phage can expand the Stx-producing population in a human or animal host, and phage infection has been shown to modulate virulence in vitro and in vivo. Acellular factors, particularly colicins and microcins, can kill O157:H7 cells but may also trigger Stx expression in the process. Colicins, microcins, and other bacteriocins have diverse cellular targets, and many such molecules remain uncharacterized. The identification of additional Stx-amplifying microbial interactions will improve our understanding of E. coli O157:H7 infections and help elucidate the intricate regulation of pathogenicity in EHEC strains.

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