scholarly journals The Tetrazole VT-1161 Is a Potent Inhibitor of Trichophyton rubrum through Its Inhibition of T. rubrum CYP51

2017 ◽  
Vol 61 (7) ◽  
Author(s):  
Andrew G. S. Warrilow ◽  
Josie E. Parker ◽  
Claire L. Price ◽  
Edward P. Garvey ◽  
William J. Hoekstra ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Trichophyton Rubrum ◽  
Potent Inhibitor ◽  
Geometric Mean ◽  
Tight Binding ◽  
Cytochrome P450s ◽  
Turnover Number ◽  
Content Type ◽  
Investigational Agent

ABSTRACT Prior to characterization of antifungal inhibitors that target CYP51, Trichophyton rubrum CYP51 was expressed in Escherichia coli, purified, and characterized. T. rubrum CYP51 bound lanosterol, obtusifoliol, and eburicol with similar affinities (dissociation constant [Kd ] values, 22.7, 20.3, and 20.9 μM, respectively) but displayed substrate specificity, insofar as only eburicol was demethylated in CYP51 reconstitution assays (turnover number, 1.55 min−1; Km value, 2 μM). The investigational agent VT-1161 bound tightly to T. rubrum CYP51 (Kd = 242 nM) with an affinity similar to that of clotrimazole, fluconazole, ketoconazole, and voriconazole (Kd values, 179, 173, 312, and 304 nM, respectively) and with an affinity lower than that of itraconazole (Kd = 53 nM). Determinations of 50% inhibitory concentrations (IC50s) using 0.5 μM CYP51 showed that VT-1161 was a tight-binding inhibitor of T. rubrum CYP51 activity, yielding an IC50 of 0.14 μM, whereas itraconazole, fluconazole, and ketoconazole had IC50s of 0.26, 0.4, and 0.6 μM, respectively. When the activity of VT-1161 was tested against 34 clinical isolates, VT-1161 was a potent inhibitor of T. rubrum growth, with MIC50, MIC90, and geometric mean MIC values of ≤0.03, 0.06, and 0.033 μg ml−1, respectively. With its selectivity versus human CYP51 and drug-metabolizing cytochrome P450s having already been established, VT-1161 should prove to be safe and effective in combating T. rubrum infections in patients.

scholarly journals Characterization of Epidemic IncI1-Iγ Plasmids Harboring Ambler Class A and C Genes in Escherichia coli and Salmonella enterica from Animals and Humans

2015 ◽  
Vol 59 (9) ◽  
pp. 5357-5365 ◽  
Author(s):  
Hilde Smith ◽  
Alex Bossers ◽  
Frank Harders ◽  
Guanghui Wu ◽  
Neil Woodford ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Salmonella Enterica ◽  
Phylogenetic Trees ◽  
Functional Study ◽  
Content Type ◽  
Gene Associations ◽  
Genes Encoding ◽  
Marked Resistance

ABSTRACTThe aim of the study was to identify the plasmid-encoded factors contributing to the emergence and spread of epidemic IncI1-Iγ plasmids obtained fromEscherichia coliandSalmonella entericaisolates from animal and human reservoirs. For this, 251 IncI1-Iγ plasmids carrying various extended-spectrum β-lactamase (ESBL) or AmpC β-lactamase genes were compared using plasmid multilocus sequence typing (pMLST). Thirty-two of these plasmids belonging to different pMLST types were sequenced using Roche 454 and Illumina platforms. Epidemic IncI1-Iγ plasmids could be assigned to various dominant clades, whereas rarely detected plasmids clustered together as a distinct clade. Similar phylogenetic trees were obtained using only the plasmid backbone sequences, showing that the differences observed between the plasmids belonging to distinct clades resulted mainly from differences between their backbone sequences. Plasmids belonging to the various clades differed particularly in the presence/absence of genes encoding partitioning and addiction systems, which contribute to stable inheritance during cell division and plasmid maintenance. Despite this, plasmids belonging to the various phylogenetic clades also showed marked resistance gene associations, indicating the circulation of successful plasmid-gene combinations. The variation intraYandexcAgenes found in IncI1-Iγ plasmids is conserved within pMLST sequence types and plays a role in incompatibility, although functional study is needed to elucidate the role of these genes in plasmid epidemiology.

scholarly journals Identification and Characterization of Genes Required for 5-Hydroxyuridine Synthesis in Bacillus subtilis and Escherichia coli tRNA

2019 ◽  
Vol 201 (20) ◽  
Author(s):  
Charles T. Lauhon
Keyword(s):  
Escherichia Coli ◽  
Bacillus Subtilis ◽  
Genomic Context ◽  
Similar Reduction ◽  
Content Type ◽  
E Coli ◽  
Wobble Position ◽  
Fertile Ground ◽  
And Function

ABSTRACT In bacteria, tRNAs that decode 4-fold degenerate family codons and have uridine at position 34 of the anticodon are typically modified with either 5-methoxyuridine (mo5U) or 5-methoxycarbonylmethoxyuridine (mcmo5U). These modifications are critical for extended recognition of some codons at the wobble position. Whereas the alkylation steps of these modifications have been described, genes required for the hydroxylation of U34 to give 5-hydroxyuridine (ho5U) remain unknown. Here, a number of genes in Escherichia coli and Bacillus subtilis are identified that are required for wild-type (wt) levels of ho5U. The yrrMNO operon is identified in B. subtilis as important for the biosynthesis of ho5U. Both yrrN and yrrO are homologs to peptidase U32 family genes, which includes the rlhA gene required for ho5C synthesis in E. coli. Deletion of either yrrN or yrrO, or both, gives a 50% reduction in mo5U tRNA levels. In E. coli, yegQ was found to be the only one of four peptidase U32 genes involved in ho5U synthesis. Interestingly, this mutant shows the same 50% reduction in (m)cmo5U as that observed for mo5U in the B. subtilis mutants. By analyzing the genomic context of yegQ homologs, the ferredoxin YfhL is shown to be required for ho5U synthesis in E. coli to the same extent as yegQ. Additional genes required for Fe-S biosynthesis and biosynthesis of prephenate give the same 50% reduction in modification. Together, these data suggest that ho5U biosynthesis in bacteria is similar to that of ho5C, but additional genes and substrates are required for complete modification. IMPORTANCE Modified nucleotides in tRNA serve to optimize both its structure and function for accurate translation of the genetic code. The biosynthesis of these modifications has been fertile ground for uncovering unique biochemistry and metabolism in cells. In this work, genes that are required for a novel anaerobic hydroxylation of uridine at the wobble position of some tRNAs are identified in both Bacillus subtilis and Escherichia coli. These genes code for Fe-S cluster proteins, and their deletion reduces the levels of the hydroxyuridine by 50% in both organisms. Additional genes required for Fe-S cluster and prephenate biosynthesis and a previously described ferredoxin gene all display a similar reduction in hydroxyuridine levels, suggesting that still other genes are required for the modification.

scholarly journals Molecular Characterization of Commensal Escherichia coli Adapted to Different Compartments of the Porcine Gastrointestinal Tract

2012 ◽  
Vol 78 (19) ◽  
pp. 6799-6803 ◽  
Author(s):  
Sam Abraham ◽  
David M. Gordon ◽  
James Chin ◽  
Huub J. M. Brouwers ◽  
Peter Njuguna ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Gastrointestinal Tract ◽  
Random Amplified Polymorphic Dna ◽  
Content Type ◽  
E Coli ◽  
Plasmid Replicon ◽  
Different Strains ◽  
Different Characteristics

ABSTRACTThe role ofEscherichia colias a pathogen has been the focus of considerable study, while much less is known about it as a commensal and how it adapts to and colonizes different environmental niches within the mammalian gut. In this study, we characterizeEscherichia coliorganisms (n= 146) isolated from different regions of the intestinal tracts of eight pigs (dueodenum, ileum, colon, and feces). The isolates were typed using the method of random amplified polymorphic DNA (RAPD) and screened for the presence of bacteriocin genes and plasmid replicon types. Molecular analysis of variance using the RAPD data showed thatE. coliisolates are nonrandomly distributed among different gut regions, and that gut region accounted for 25% (P< 0.001) of the observed variation among strains. Bacteriocin screening revealed that a bacteriocin gene was detected in 45% of the isolates, with 43% carrying colicin genes and 3% carrying microcin genes. Of the bacteriocins observed (H47, E3, E1, E2, E7, Ia/Ib, and B/M), the frequency with which they were detected varied with respect to gut region for the colicins E2, E7, Ia/Ib, and B/M. The plasmid replicon typing gave rise to 25 profiles from the 13 Inc types detected. Inc F types were detected most frequently, followed by Inc HI1 and N types. Of the Inc types detected, 7 were nonrandomly distributed among isolates from the different regions of the gut. The results of this study indicate that not only may the different regions of the gastrointestinal tract harbor different strains ofE. colibut also that strains from different regions have different characteristics.

scholarly journals Characterization of blaCTX-M-27/F1:A2:B20 Plasmids Harbored by Escherichia coli Sequence Type 131 Sublineage C1/H30R Isolates Spreading among Elderly Japanese in Nonacute-Care Settings

2020 ◽  
Vol 64 (5) ◽  
Author(s):  
Nao Matsuo ◽  
Rina Nonogaki ◽  
Michiko Hayashi ◽  
Jun-ichi Wachino ◽  
Masahiro Suzuki ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Care Facility ◽  
Sequence Type ◽  
Health Concern ◽  
Term Care ◽  
Content Type ◽  
Long Term Care Facility ◽  
Elderly Japanese

ABSTRACT We characterized 29 blaCTX-M-27-harboring plasmids of Escherichia coli sequence type 131 (ST131) sublineage C1/H30R isolates from healthy individuals and long-term-care facility (LTCF) residents. Most (27/29) plasmids were of the FIA, FIB, and FII multireplicon type with the same plasmid multilocus sequence typing (pMLST). Several plasmids (7/23) from LTCF residents harbored only blaCTX-M-27 as the resistance gene; however, their fundamental structures were very similar to those of previously isolated blaCTX-M-27/F1:A2:B20 plasmids, suggesting their prevalence as a newly arising public health concern.

scholarly journals Characterization of Multidrug-Resistant Escherichia coli Isolates from Animals Presenting at a University Veterinary Hospital

2011 ◽  
Vol 77 (20) ◽  
pp. 7104-7112 ◽  
Author(s):  
Maria Karczmarczyk ◽  
Yvonne Abbott ◽  
Ciara Walsh ◽  
Nola Leonard ◽  
Séamus Fanning
Keyword(s):  
Escherichia Coli ◽  
Molecular Mechanisms ◽  
Gene Array ◽  
Multidrug Resistant ◽  
Genetic Determinants ◽  
Gene Encoding ◽  
Content Type ◽  
E Coli ◽  
Class 1

ABSTRACTIn this study, we examined molecular mechanisms associated with multidrug resistance (MDR) in a collection ofEscherichia coliisolates recovered from hospitalized animals in Ireland. PCR and DNA sequencing were used to identify genes associated with resistance. Class 1 integrons were prevalent (94.6%) and contained gene cassettes recognized previously and implicated mainly in resistance to aminoglycosides, β-lactams, and trimethoprim (aadA1,dfrA1-aadA1,dfrA17-aadA5,dfrA12-orfF-aadA2,blaOXA-30-aadA1,aacC1-orf1-orf2-aadA1,dfr7). Class 2 integrons (13.5%) contained thedfrA1-sat1-aadA1gene array. The most frequently occurring phenotypes included resistance to ampicillin (97.3%), chloramphenicol (75.4%), florfenicol (40.5%), gentamicin (54%), neomycin (43.2%), streptomycin (97.3%), sulfonamide (98.6%), and tetracycline (100%). The associated resistance determinants detected includedblaTEM,cat,floR,aadB,aphA1,strA-strB,sul2, andtet(B), respectively. TheblaCTX-M-2gene, encoding an extended-spectrum β-lactamase (ESβL), andblaCMY-2, encoding an AmpC-like enzyme, were identified in 8 and 18 isolates, respectively. The mobility of the resistance genes was demonstrated using conjugation assays with a representative selection of isolates. High-molecular-weight plasmids were found to be responsible for resistance to multiple antimicrobial compounds. The study demonstrated that animal-associated commensalE. coliisolates possess a diverse repertoire of transferable genetic determinants. Emergence of ESβLs and AmpC-like enzymes is particularly significant. To our knowledge, theblaCTX-M-2gene has not previously been reported in Ireland.

scholarly journals Characterization of the New AmpC β-Lactamase FOX-8 Reveals a Single Mutation, Phe313Leu, Located in the R2 Loop That Affects Ceftazidime Hydrolysis

2013 ◽  
Vol 57 (10) ◽  
pp. 5158-5161 ◽  
Author(s):  
Francisco José Pérez-Llarena ◽  
Frédéric Kerff ◽  
Laura Zamorano ◽  
María Carmen Fernández ◽  
Maria Luz Nuñez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Kinetic Analysis ◽  
Clinical Strain ◽  
Single Mutation ◽  
Content Type ◽  
E Coli ◽  
Fold Reduction ◽  
Class C

ABSTRACTA novel class C β-lactamase (FOX-8) was isolated from a clinical strain ofEscherichia coli. The FOX-8 enzyme possessed a unique substitution (Phe313Leu) compared to FOX-3. IsogenicE. colistrains carrying FOX-8 showed an 8-fold reduction in resistance to ceftazidime relative to FOX-3. In a kinetic analysis, FOX-8 displayed a 33-fold reduction inkcat/Kmfor ceftazidime compared to FOX-3. In the FOX family of β-lactamases, the Phe313 residue located in the R2 loop affects ceftazidime hydrolysis and alters the phenotype ofE. colistrains carrying this variant.

scholarly journals Characterization of SXT/R391 Integrative and Conjugative Elements in Proteus mirabilis Isolates from Food-Producing Animals in China

2016 ◽  
Vol 60 (3) ◽  
pp. 1935-1938 ◽  
Author(s):  
Chang-Wei Lei ◽  
An-Yun Zhang ◽  
Hong-Ning Wang ◽  
Bi-Hui Liu ◽  
Li-Qin Yang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Resistance Genes ◽  
Proteus Mirabilis ◽  
Whole Genome ◽  
Content Type ◽  
Integrative And Conjugative Elements ◽  
Animal Farms

SXT/R391 integrative and conjugative elements (ICEs) were detected in 8 out of 125Proteus mirabilisisolates from food-producing animals in China. Whole-genome sequencing revealed that seven ICEs were identical to ICEPmiJpn1, carrying the cephalosporinase geneblaCMY-2. Another one, designated ICEPmiChn1, carried five resistance genes. All eight ICEs could be transferred toEscherichia colivia conjugation. The results highlight the idea that animal farms are important reservoir of the SXT/R391 ICE-containingP. mirabilis.

scholarly journals Characterization of a Novel Pyranopyridine Inhibitor of the AcrAB Efflux Pump of Escherichia coli

2013 ◽  
Vol 58 (2) ◽  
pp. 722-733 ◽  
Author(s):  
Timothy J. Opperman ◽  
Steven M. Kwasny ◽  
Hong-Suk Kim ◽  
Son T. Nguyen ◽  
Chad Houseweart ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Efflux Pump ◽  
Efflux Pumps ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Resistance Nodulation Division ◽  
Rnd Efflux Pumps ◽  
Acrab Efflux Pump

ABSTRACTMembers of the resistance-nodulation-division (RND) family of efflux pumps, such as AcrAB-TolC ofEscherichia coli, play major roles in multidrug resistance (MDR) in Gram-negative bacteria. A strategy for combating MDR is to develop efflux pump inhibitors (EPIs) for use in combination with an antibacterial agent. Here, we describe MBX2319, a novel pyranopyridine EPI with potent activity against RND efflux pumps of theEnterobacteriaceae. MBX2319 decreased the MICs of ciprofloxacin (CIP), levofloxacin, and piperacillin versusE. coliAB1157 by 2-, 4-, and 8-fold, respectively, but did not exhibit antibacterial activity alone and was not active against AcrAB-TolC-deficient strains. MBX2319 (3.13 μM) in combination with 0.016 μg/ml CIP (minimally bactericidal) decreased the viability (CFU/ml) ofE. coliAB1157 by 10,000-fold after 4 h of exposure, in comparison with 0.016 μg/ml CIP alone. In contrast, phenyl-arginine-β-naphthylamide (PAβN), a known EPI, did not increase the bactericidal activity of 0.016 μg/ml CIP at concentrations as high as 100 μM. MBX2319 increased intracellular accumulation of the fluorescent dye Hoechst 33342 in wild-type but not AcrAB-TolC-deficient strains and did not perturb the transmembrane proton gradient. MBX2319 was broadly active againstEnterobacteriaceaespecies andPseudomonas aeruginosa. MBX2319 is a potent EPI with possible utility as an adjunctive therapeutic agent for the treatment of infections caused by Gram-negative pathogens.

scholarly journals Characterization of CIA-1, an Ambler Class A Extended-Spectrum β-Lactamase from Chryseobacterium indologenes

2011 ◽  
Vol 56 (1) ◽  
pp. 588-590 ◽  
Author(s):  
Takehisa Matsumoto ◽  
Mika Nagata ◽  
Nau Ishimine ◽  
Kenji Kawasaki ◽  
Kazuyoshi Yamauchi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Reference Strain ◽  
Content Type ◽  
Chryseobacterium Indologenes ◽  
Esbl Gene ◽  
Class A ◽  
Extended Spectrum ◽  
Class B ◽  
Lactamase Gene

ABSTRACTAn Ambler class A β-lactamase gene,blaCIA-1, was cloned from the reference strainChryseobacterium indologenesATCC 29897 and expressed inEscherichia coliBL21. TheblaCIA-1gene encodes a novel extended-spectrum β-lactamase (ESBL) that shared 68% and 60% identities with the CGA-1 and CME-1 β-lactamases, respectively.blaCIA-1-like genes were detected from clinical isolates. In addition to the metallo-β-lactamase IND of Ambler class B,C. indologeneshas a class A ESBL gene,blaCIA-1, located on the chromosome.

scholarly journals Identification and Biochemical Characterization of a Novel Protein Phosphatase 2C-Like Ser/Thr Phosphatase in Escherichia coli

2018 ◽  
Vol 200 (18) ◽  
Author(s):  
Krithika Rajagopalan ◽  
Elizabeth Nagle ◽  
Jonathan Dworkin
Keyword(s):  
Escherichia Coli ◽  
Protein Phosphatase ◽  
Biochemical Characterization ◽  
Regulatory Protein ◽  
Negative Bacterium ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Novel Protein

Regulatory protein phosphorylation is a conserved mechanism of signaling in all biological systems. Recent phosphoproteomic analyses of phylogenetically diverse bacteria, including the model Gram-negative bacteriumEscherichia coli, demonstrate that many proteins are phosphorylated on serine or threonine residues. In contrast to phosphorylation on histidine or aspartate residues, phosphorylation of serine and threonine residues is stable and requires the action of a partner Ser/Thr phosphatase to remove the modification. Although a number of Ser/Thr kinases have been reported inE. coli, no partner Ser/Thr phosphatases have been identified. Here, we biochemically characterize a novel Ser/Thr phosphatase that acts to dephosphorylate a Ser/Thr kinase that is encoded in the same operon.

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