scholarly journals Lsr operon is associated with AI-2 transfer and pathogenicity in avian pathogenic Escherichia coli

2019 ◽  
Vol 50 (1) ◽  
Author(s):  
Jiakun Zuo ◽  
Huifang Yin ◽  
Jiangang Hu ◽  
Jinfeng Miao ◽  
Zhaoguo Chen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Wild Type Strain ◽  
Binding Capacity ◽  
Bacterial Load ◽  
Transcriptional Analysis ◽  
Wild Type ◽  
Signal Molecule ◽  
Autoinducer 2

AbstractThe function of Autoinducer-2 (AI-2) which acts as the signal molecule of LuxS-mediated quorum sensing, is regulated through the lsr operon (which includes eight genes: lsrK, lsrR, lsrA, lsrC, lsrD, lsrB, lsrF, and lsrG). However, the functions of the lsr operon remain unclear in avian pathogenic Escherichia coli (APEC), which causes severe respiratory and systemic diseases in poultry. In this study, the presence of the lsr operon in 60 APEC clinical strains (serotypes O1, O2, and O78) was investigated and found to be correlated with serotype and has the highest detection rate in O78. The AI-2 binding capacity of recombinant protein LsrB of APEC (APEC-LsrB) was verified and was found to bind to AI-2 in vitro. In addition, the lsr operon was mutated in an APEC strain (APEC94Δlsr(Cm)) and the mutant was found to be defective in motility and AI-2 uptake. Furthermore, deletion of the lsr operon attenuated the virulence of APEC, with the LD50 of APEC94Δlsr(Cm) decreasing 294-fold compared with wild-type strain APEC94. The bacterial load in the blood, liver, spleen, and kidneys of ducks infected with APEC94Δlsr(Cm) decreased significantly (p < 0.0001). The results of transcriptional analysis showed that 62 genes were up-regulated and 415 genes were down-regulated in APEC94Δlsr(Cm) compared with the wild-type strain and some of the down-regulated genes were associated with the virulence of APEC. In conclusion, our study suggests that lsr operon plays a role in the pathogenesis of APEC.

scholarly journals Identification of Concomitant Infection with Chlamydia trachomatis IncA-Negative Mutant and Wild-Type Strains by Genomic, Transcriptional, and Biological Characterizations

2008 ◽  
Vol 76 (12) ◽  
pp. 5438-5446 ◽  
Author(s):  
Robert J. Suchland ◽  
Brendan M. Jeffrey ◽  
Minsheng Xia ◽  
Ajay Bhatia ◽  
Hencelyn G. Chu ◽  
...  
Keyword(s):  
Chlamydia Trachomatis ◽  
Genome Sequence ◽  
Type Strain ◽  
Wild Type Strain ◽  
Genomic Sequence ◽  
Genomic Structure ◽  
Transcriptional Analysis ◽  
Clinical Samples ◽  
Wild Type

ABSTRACT Clinical isolates of Chlamydia trachomatis that lack IncA on their inclusion membrane form nonfusogenic inclusions and have been associated with milder, subclinical infections in patients. The molecular events associated with the generation of IncA-negative strains and their roles in chlamydial sexually transmitted infections are not clear. We explored the biology of the IncA-negative strains by analyzing their genomic structure, transcription, and growth characteristics in vitro and in vivo in comparison with IncA-positive C. trachomatis strains. Three clinical samples were identified that contained a mixture of IncA-positive and -negative same-serovar C. trachomatis populations, and two more such pairs were found in serial isolates from persistently infected individuals. Genomic sequence analysis of individual strains from each of two serovar-matched pairs showed that these pairs were very similar genetically. In contrast, the genome sequence of an unmatched IncA-negative strain contained over 5,000 nucleotide polymorphisms relative to the genome sequence of a serovar-matched but otherwise unlinked strain. Transcriptional analysis, in vitro culture kinetics, and animal modeling demonstrated that IncA-negative strains isolated in the presence of a serovar-matched wild-type strain are phenotypically more similar to the wild-type strain than are IncA-negative strains isolated in the absence of a serovar-matched wild-type strain. These studies support a model suggesting that a change from an IncA-positive strain to the previously described IncA-negative phenotype may involve multiple steps, the first of which involves a translational inactivation of incA, associated with subsequent unidentified steps that lead to the observed decrease in transcript level, differences in growth rate, and differences in mouse infectivity.

scholarly journals Finding Regulators Associated with the Expression of the Long Polar Fimbriae in Enteropathogenic Escherichia coli

2015 ◽  
Vol 197 (23) ◽  
pp. 3658-3665 ◽  
Author(s):  
Jia Hu ◽  
Brittany N. Ross ◽  
Roberto J. Cieza ◽  
Alfredo G. Torres
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Content Type ◽  
E Coli ◽  
Colonization Factor ◽  
Initial Adhesion ◽  
Intestinal Adhesion

ABSTRACTEnteropathogenicEscherichia coli(EPEC) is a human pathogen that requires initial adhesion to the intestine in order to cause disease. Multiple adhesion factors have been identified inE. colistrains, among them the long polar fimbriae (Lpf), a colonization factor associated with intestinal adhesion. The conditions of Lpf expression are well understood in enterohemorrhagicE. coli(EHEC); however, the expression of EPEClpfhas been found to be repressed under anyin vitrocondition tested. Therefore, we decided to identify those factors silencing expression of EPEClpf. Because histone-like nucleoid structuring protein (H-NS) is a known repressor of EHEClpf, we tested it and found that H-NS is a repressor of EPEClpf. We also found that the adhesion of the EPEC Δhnsstrain was significantly enhanced compared to the wild-type strain. Becauselpfexpression was modestly increased in thehnsmutant, transposon mutagenesis was performed to find a strain displaying higherlpfexpression than EPEC Δhns. One Tn5insertion was identified within theyhjXgene, and furtherin vitrocharacterization revealed increasedlpfexpression and adhesion to Caco-2 cells compared with EPEC Δhns. However, in a murine model of intestinal infection, the EPEC Δhnsand EPEC ΔhnsTn5mutants had only a slight change in colonization pattern compared to the wild-type strain. Our data showed that EPEC Lpf is transcribed, but its role in EPEC intestinal colonization requires further analysis.IMPORTANCEData are presented demonstrating that the long polar fimbriae (lpf) operon in enteropathogenicE. coli(EPEC) is highly regulated; however, derepression occurs by mutagenizing two proteins associated with its control. The study demonstrates that the EPEClpfoperon can be expressed and, therefore, participates in the EPEC adherence phenotype.

scholarly journals Autoinducer 2 Controls Biofilm Formation in Escherichia coli through a Novel Motility Quorum-Sensing Regulator (MqsR, B3022)

2006 ◽  
Vol 188 (1) ◽  
pp. 305-316 ◽  
Author(s):  
Andrés F. González Barrios ◽  
Rongjun Zuo ◽  
Yoshifumi Hashimoto ◽  
Li Yang ◽  
William E. Bentley ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Continuous Flow ◽  
Biofilm Formation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Wild Type ◽  
Flow Cells ◽  
Autoinducer 2 ◽  
K 12

ABSTRACT The cross-species bacterial communication signal autoinducer 2 (AI-2), produced by the purified enzymes Pfs (nucleosidase) and LuxS (terminal synthase) from S-adenosylhomocysteine, directly increased Escherichia coli biofilm mass 30-fold. Continuous-flow cells coupled with confocal microscopy corroborated these results by showing the addition of AI-2 significantly increased both biofilm mass and thickness and reduced the interstitial space between microcolonies. As expected, the addition of AI-2 to cells which lack the ability to transport AI-2 (lsr null mutant) failed to stimulate biofilm formation. Since the addition of AI-2 increased cell motility through enhanced transcription of five motility genes, we propose that AI-2 stimulates biofilm formation and alters its architecture by stimulating flagellar motion and motility. It was also found that the uncharacterized protein B3022 regulates this AI-2-mediated motility and biofilm phenotype through the two-component motility regulatory system QseBC. Deletion of b3022 abolished motility, which was restored by expressing b3022 in trans. Deletion of b3022 also decreased biofilm formation significantly, relative to the wild-type strain in three media (46 to 74%) in 96-well plates, as well as decreased biomass (8-fold) and substratum coverage (19-fold) in continuous-flow cells with minimal medium (growth rate not altered and biofilm restored by expressing b3022 in trans). Deleting b3022 changed the wild-type biofilm architecture from a thick (54-μm) complex structure to one that contained only a few microcolonies. B3022 positively regulates expression of qseBC, flhD, fliA, and motA, since deleting b3022 decreased their transcription by 61-, 25-, 2.4-, and 18-fold, respectively. Transcriptome analysis also revealed that B3022 induces crl (26-fold) and flhCD (8- to 27-fold). Adding AI-2 (6.4 μM) increased biofilm formation of wild-type K-12 MG1655 but not that of the isogenic b3022, qseBC, fliA, and motA mutants. Adding AI-2 also increased motA transcription for the wild-type strain but did not stimulate motA transcription for the b3022 and qseB mutants. Together, these results indicate AI-2 induces biofilm formation in E. coli through B3022, which then regulates QseBC and motility; hence, b3022 has been renamed the motility quorum-sensing regulator gene (the mqsR gene).

scholarly journals Quorum Sensing Is a Global Regulatory Mechanism in Enterohemorrhagic Escherichia coli O157:H7

2001 ◽  
Vol 183 (17) ◽  
pp. 5187-5197 ◽  
Author(s):  
Vanessa Sperandio ◽  
Alfredo G. Torres ◽  
Jorge A. Girón ◽  
James B. Kaper
Keyword(s):  
Escherichia Coli ◽  
Quorum Sensing ◽  
Type Strain ◽  
Wild Type Strain ◽  
Regulatory Mechanism ◽  
Sos Response ◽  
Enterohemorrhagic Escherichia Coli ◽  
Trna Genes ◽  
Wild Type ◽  
E Coli

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is responsible for outbreaks of bloody diarrhea and hemolytic-uremic syndrome in many countries. EHEC virulence mechanisms include the production of Shiga toxins (Stx) and formation of attaching and effacing (AE) lesions on intestinal epithelial cells. We recently reported that genes involved in the formation of the AE lesion were regulated by quorum sensing through autoinducer-2, which is synthesized by the product of the luxS gene. In this study we hybridized an E. coli gene array with cDNA synthesized from RNA that was extracted from EHEC strain 86-24 and its isogenicluxS mutant. We observed that 404 genes were regulated by luxS at least fivefold, which comprises approximately 10% of the array genes; 235 of these genes were up-regulated and 169 were down-regulated in the wild-type strain compared to in theluxS mutant. Down-regulated genes included several involved in cell division, as well as ribosomal and tRNA genes. Consistent with this pattern of gene expression, theluxS mutant grows faster than the wild-type strain (generation times of 37.5 and 60 min, respectively, in Dulbecco modified Eagle medium). Up-regulated genes included several involved in the expression and assembly of flagella, motility, and chemotaxis. Using operon::lacZ fusions to class I, II, and III flagellar genes, we were able to confirm this transcriptional regulation. We also observed fewer flagella by Western blotting and electron microscopy and decreased motility halos in semisolid agar in the luxS mutant. The average swimming speeds for the wild-type strain and the luxS mutant are 12.5 and 6.6 μm/s, respectively. We also observed an increase in the production of Stx due to quorum sensing. Genes encoding Stx, which are transcribed along with λ-like phage genes, are induced by an SOS response, and genes involved in the SOS response were also regulated by quorum sensing. These results indicate that quorum sensing is a global regulatory mechanism for basic physiological functions of E. coli as well as for virulence factors.

scholarly journals Ionophore activity of sarcotoxin I, a bactericidal protein of Sarcophaga peregrina

1985 ◽  
Vol 229 (2) ◽  
pp. 453-458 ◽  
Author(s):  
M Okada ◽  
S Natori
Keyword(s):  
Escherichia Coli ◽  
Oxidative Phosphorylation ◽  
Type Strain ◽  
Wild Type Strain ◽  
Bactericidal Effect ◽  
Proton Gradient ◽  
Wild Type ◽  
Atp Pool

When Escherichia coli was treated with sarcotoxin I, a potent bactericidal protein of Sarcophaga peregrina (fleshfly), K+ inside of the cells leaked out rapidly and the ATP pool of the cells rapidly decreased. These results suggested that the bactericidal effect of sarcotoxin I was due to its ionophore activity, and that it blocked the generation of ATP by inhibiting formation of the proton gradient essential for oxidative phosphorylation. This was confirmed by use of an uncA mutant, which was much less susceptible than the wild-type strain to sarcotoxin I under fixed ionic conditions.

scholarly journals Accumulation of Inorganic Polyphosphate in phoU Mutants of Escherichia coli and Synechocystis sp. Strain PCC6803

2002 ◽  
Vol 68 (8) ◽  
pp. 4107-4110 ◽  
Author(s):  
Tomohiro Morohoshi ◽  
Tatsuya Maruo ◽  
Yoko Shirai ◽  
Junichi Kato ◽  
Tsukasa Ikeda ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Wild Type Strain ◽  
Biological Process ◽  
Negative Regulator ◽  
Wild Type ◽  
Inorganic Polyphosphate ◽  
Insertional Inactivation ◽  
Synechocystis Sp ◽  
Polyp Accumulation

ABSTRACT The biological process for phosphate (Pi) removal is based on the use of bacteria capable of accumulating inorganic polyphosphate (polyP). We obtained Escherichia coli mutants which accumulate a large amount of polyP. The polyP accumulation in these mutants was ascribed to a mutation of the phoU gene that encodes a negative regulator of the Pi regulon. Insertional inactivation of the phoU gene also elevated the intracellular level of polyP in Synechocystis sp. strain PCC6803. The mutant could remove fourfold more Pi from the medium than the wild-type strain removed.

scholarly journals Construction and Characterization of aMycobacterium tuberculosis Mutant Lacking the Alternate Sigma Factor Gene, sigF

2000 ◽  
Vol 68 (10) ◽  
pp. 5575-5580 ◽  
Author(s):  
Ping Chen ◽  
Rafael E. Ruiz ◽  
Qing Li ◽  
Richard F. Silver ◽  
William R. Bishai
Keyword(s):  
Stationary Phase ◽  
Sigma Factor ◽  
Type Strain ◽  
Wild Type Strain ◽  
Axenic Culture ◽  
Wild Type ◽  
Intracellular Growth ◽  
Factor Gene ◽  
Time To Death

ABSTRACT The alternate RNA polymerase sigma factor gene, sigF, which is expressed in stationary phase and under stress conditions in vitro, has been deleted in the virulent CDC1551 strain ofMycobacterium tuberculosis. The growth rate of the ΔsigF mutant was identical to that of the isogenic wild-type strain in exponential phase, although in stationary phase the mutant achieved a higher density than the wild type. The mutant showed increased susceptibility to rifampin and rifapentine. Additionally, the ΔsigF mutant displayed diminished uptake of chenodeoxycholate, and this effect was reversed by complementation with a wild-type sigF gene. No differences in short-term intracellular growth between mutant and wild-type organisms within human monocytes were observed. Similarly, the organisms did not differ in their susceptibilities to lymphocyte-mediated inhibition of intracellular growth. However, mice infected with the ΔsigF mutant showed a median time to death of 246 days compared with 161 days for wild-type strain-infected animals (P < 0.001). These data indicate that M. tuberculosis sigF is a nonessential alternate sigma factor both in axenic culture and for survival in macrophages in vitro. While the ΔsigF mutant produces a lethal infection of mice, it is less virulent than its wild-type counterpart by time-to-death analysis.

scholarly journals Mutations Conferring Aminoglycoside and Spectinomycin Resistance in Borrelia burgdorferi

2006 ◽  
Vol 50 (2) ◽  
pp. 445-452 ◽  
Author(s):  
Daniel Criswell ◽  
Virginia L. Tobiason ◽  
J. Stephen Lodmell ◽  
D. Scott Samuels
Keyword(s):  
16S Rrna ◽  
Borrelia Burgdorferi ◽  
Type Strain ◽  
Wild Type Strain ◽  
Small Subunit ◽  
Wild Type ◽  
Spectinomycin Resistance ◽  
E Coli ◽  
Resistant Mutants

ABSTRACT We have isolated and characterized in vitro mutants of the Lyme disease agent Borrelia burgdorferi that are resistant to spectinomycin, kanamycin, gentamicin, or streptomycin, antibiotics that target the small subunit of the ribosome. 16S rRNA mutations A1185G and C1186U, homologous to Escherichia coli nucleotides A1191 and C1192, conferred >2,200-fold and 1,300-fold resistance to spectinomycin, respectively. A 16S rRNA A1402G mutation, homologous to E. coli A1408, conferred >90-fold resistance to kanamycin and >240-fold resistance to gentamicin. Two mutations were identified in the gene for ribosomal protein S12, at a site homologous to E. coli residue Lys-87, in mutants selected in streptomycin. Substitutions at codon 88, K88R and K88E, conferred 7-fold resistance and 10-fold resistance, respectively, to streptomycin on B. burgdorferi. The 16S rRNA A1185G and C1186U mutations, associated with spectinomycin resistance, appeared in a population of B. burgdorferi parental strain B31 at a high frequency of 6 × 10−6. These spectinomycin-resistant mutants successfully competed with the wild-type strain during 100 generations of coculture in vitro. The aminoglycoside-resistant mutants appeared at a frequency of 3 × 10−9 to 1 ×10−7 in a population and were unable to compete with wild-type strain B31 after 100 generations. This is the first description of mutations in the B. burgdorferi ribosome that confer resistance to antibiotics. These results have implications for the evolution of antibiotic resistance, because the 16S rRNA mutations conferring spectinomycin resistance have no significant fitness cost in vitro, and for the development of new selectable markers.

scholarly journals SdiA Aids Enterohemorrhagic Escherichia coli Carriage by Cattle Fed a Forage or Grain Diet

2013 ◽  
Vol 81 (9) ◽  
pp. 3472-3478 ◽  
Author(s):  
Haiqing Sheng ◽  
Y. N. Nguyen ◽  
Carolyn J. Hovde ◽  
Vanessa Sperandio
Keyword(s):  
Escherichia Coli ◽  
Type Strain ◽  
Deletion Mutant ◽  
Wild Type Strain ◽  
Rumen Fluid ◽  
Acid Resistance ◽  
Enterohemorrhagic Escherichia Coli ◽  
Dependent Manner ◽  
Bacterial Survival ◽  
Wild Type

ABSTRACTEnterohemorrhagicEscherichia coli(EHEC) causes hemorrhagic colitis and life-threatening complications. The main reservoirs for EHEC are healthy ruminants. We reported that SdiA senses acyl homoserine lactones (AHLs) in the bovine rumen to activate expression of the glutamate acid resistance (gad) genes priming EHEC's acid resistance before they pass into the acidic abomasum. Conversely, SdiA represses expression of the locus of enterocyte effacement (LEE) genes, whose expression is not required for bacterial survival in the rumen but is necessary for efficient colonization at the rectoanal junction (RAJ) mucosa. Our previous studies show that SdiA-dependent regulation was necessary for efficient EHEC colonization of cattle fed a grain diet. Here, we compared the SdiA role in EHEC colonization of cattle fed a forage hay diet. We detected AHLs in the rumen of cattle fed a hay diet, and these AHLs activatedgadgene expression in an SdiA-dependent manner. The rumen fluid and fecal samples from hay-fed cattle were near neutrality, while the same digesta samples from grain-fed animals were acidic. Cattle fed either grain or hay and challenged with EHEC orally carried the bacteria similarly. EHEC was cleared from the rumen within days and from the RAJ mucosa after approximately one month. In competition trials, where animals were challenged with both wild-type and SdiA deletion mutant bacteria, diet did not affect the outcome that the wild-type strain was better able to persist and colonize. However, the wild-type strain had a greater advantage over the SdiA deletion mutant at the RAJ mucosa among cattle fed the grain diet.

scholarly journals Plasmid DNA Production in Proteome-Reduced Escherichia coli

2020 ◽  
Vol 8 (9) ◽  
pp. 1444
Author(s):  
Mitzi de la Cruz ◽  
Elisa A. Ramírez ◽  
Juan-Carlos Sigala ◽  
José Utrilla ◽  
Alvaro R. Lara
Keyword(s):  
Escherichia Coli ◽  
Plasmid Dna ◽  
Type Strain ◽  
Wild Type Strain ◽  
The Novel ◽  
Wild Type ◽  
Batch Mode ◽  
Cell Factories ◽  
Starting Point ◽  
In The Wild

The design of optimal cell factories requires engineering resource allocation for maximizing product synthesis. A recently developed method to maximize the saving in cell resources released 0.5% of the proteome of Escherichia coli by deleting only three transcription factors. We assessed the capacity for plasmid DNA (pDNA) production in the proteome-reduced strain in a mineral medium, lysogeny, and terrific broths. In all three cases, the pDNA yield from biomass was between 33 and 53% higher in the proteome-reduced than in its wild type strain. When cultured in fed-batch mode in shake-flask, the proteome-reduced strain produced 74.8 mg L−1 pDNA, which was four times greater than its wild-type strain. Nevertheless, the pDNA supercoiled fraction was less than 60% in all cases. Deletion of recA increased the pDNA yields in the wild type, but not in the proteome-reduced strain. Furthermore, recA mutants produced a higher fraction of supercoiled pDNA, compared to their parents. These results show that the novel proteome reduction approach is a promising starting point for the design of improved pDNA production hosts.

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