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1.  Fluorescence-Based Rapid Detection of Microbiological Contaminants in Water Samples 
The Scientific World Journal  2012;2012:234858.
Microbiological contamination of process waters is a current issue for pharmaceutical industries. Traditional methods require several days to obtain results; therefore, rapid microbiological methods are widely requested to shorten time-to-result. Milliflex Quantum was developed for the rapid detection and enumeration of microorganisms in filterable samples. It combines membrane filtration to universal fluorescent staining of viable microorganisms. This new alternative method was validated using European and United States Pharmacopeia definitions, with sterile water and/or sterile water artificially contaminated with microorganisms. The Milliflex Quantum method was demonstrated to be reliable, robust, specific, accurate, and linear over the whole range of assays following these guidelines. The Milliflex Quantum system was challenged to detect natural contaminants in different types of pharmaceutical purified process waters. Milliflex Quantum was demonstrated to detect accurately contaminants 3- to 7-fold faster than traditional membrane filtration method. The staining procedure is nondestructive allowing downstream identification following a positive result. The Milliflex Quantum offers a fast, sensitive, and robust alternative to the compendial membrane filtration method.
PMCID: PMC3353274  PMID: 22623887
2.  The AcrAB-TolC Pump Is Involved in Macrolide Resistance but Not in Telithromycin Efflux in Enterobacter aerogenes and Escherichia coli 
The role of the AcrAB-TolC pump in macrolide and ketolide susceptibility in Escherichia coli and Enterobacter aerogenes was studied. Efflux pump inhibitor restored erythromycin, clarithromycin, and telithromycin susceptibilities to multidrug-resistant isolates. No modification of telithromycin accumulation was detected in E. aerogenes acrAB or tolC derivatives compared to that in the parental strain. Two independent efflux pumps, inhibited by phenylalanine arginine β-naphthylamide, expel macrolides and telithromycin in E. aerogenes.
PMCID: PMC514773  PMID: 15328143
3.  RamA Is an Alternate Activator of the Multidrug Resistance Cascade in Enterobacter aerogenes 
Multidrug resistance (MDR) in Enterobacter aerogenes can be mediated by induction of MarA, which is triggered by certain antibiotics and phenolic compounds. In this study, we identified the gene encoding RamA, a 113-amino-acid regulatory protein belonging to the AraC-XylS transcriptional activator family, in the Enterobacter aerogenes ATCC 13048 type strain and in a clinical multiresistant isolate. Overexpression of RamA induced an MDR phenotype in drug-susceptible Escherichia coli JM109 and E. aerogenes ATCC 13048, as demonstrated by 2- to 16-fold-increased resistance to β-lactams, tetracycline, chloramphenicol, and quinolones, a decrease in porin production, and increased production of AcrA, a component of the AcrAB-TolC drug efflux pump. We show that RamA enhances the transcription of the marRAB operon but is also able to induce an MDR phenotype in a mar-deleted strain. We demonstrate here that RamA is a transcriptional activator of the Mar regulon and is also a self-governing activator of the MDR cascade.
PMCID: PMC434192  PMID: 15215103
4.  Modification of Outer Membrane Protein Profile and Evidence Suggesting an Active Drug Pump in Enterobacter aerogenes Clinical Strains 
Two clinical strains of Enterobacter aerogenes that exhibited phenotypes of multiresistance to β-lactam antibiotics, fluoroquinolones, chloramphenicol, tetracycline, and kanamycin were investigated. Both strains showed a porin pattern different from that of a susceptible strain, with a drastic reduction in the amount of the major porin but with an apparently conserved normal structure (size and immunogenicity), together with overproduction of two known outer membrane proteins, OmpX and LamB. In addition, the full-length O-polysaccharide phenotype was replaced by a semirough Ra phenotype. Moreover, in one isolate the intracellular accumulation of chloramphenicol was increased in the presence of the energy uncoupler carbonyl cyanide m-chlorophenylhydrazone, suggesting an energy-dependent efflux of chloramphenicol in this strain. The resistance strategies used by these isolates appear to be similar to that induced by stress in Escherichia coli cells.
PMCID: PMC153306  PMID: 12709321
5.  mar Operon Involved in Multidrug Resistance of Enterobacter aerogenes 
We determined the sequence of the entire marRAB operon in Enterobacter aerogenes. It is functionally and structurally analogous to the Escherichia coli operon. The overexpression of E. aerogenes MarA induces a multidrug resistance phenotype in a susceptible strain, demonstrated by a noticeable resistance to various antibiotics, a decrease in immunodetected porins, and active efflux of norfloxacin.
PMCID: PMC127096  PMID: 11897595

Results 1-5 (5)