Evaluations of the Effects of Extremely Low-Frequency Electromagnetic Fields on Growth and Antibiotic Susceptibility of Escherichia coli and Pseudomonas aeruginosa.

We aimed to investigate the effects of exposure to extremely low-frequency electromagnetic fields (2 mT; 50 Hz) on the growth rate and antibiotic sensitivity of E. coli ATCC 25922 and P. aeruginosa ATCC 27853. The electromagnetic field treatment significantly influenced the growth rate of both strains when incubated in the presence of subinhibitory concentrations of kanamycin (1 µg/mL) and amikacin (0.5 µg/mL), respectively. In particular, at 4, 6, and 8 h of incubation the number of cells was significantly decreased in bacteria exposed to electromagnetic field when compared with the control. Additionally, at 24 h of incubation, the percentage of cells increased (P. aeruginosa∼42%; E. coli∼5%) in treated groups with respect to control groups suggesting a progressive adaptive response. By contrast, no remarkable differences were found in the antibiotic susceptibility and on the growth rate of both bacteria comparing exposed groups with control groups.

In vitro selection and characterization of mutants in TEM-1-producing Escherichia coli by ceftazidime and ceftibuten.

The present work was undertaken to study the ability of ceftazidime and ceftibuten to selectin vitro Escherichia coli HB101 harboring bla(TEM-1) beta-lactamase gene. Minimum inhibitory concentrations (MICs) of ceftazidime and ceftibuten were increased by a factor of 32, overcoming in the case of ceftazidime the breakpoint for clinical resistance. Outer membrane protein analysis and PCR for bla(TEM )alleles revealed that ceftazidime and ceftibuten select for different resistance mechanisms. Ceftazidime created mutants that encode an extended-spectrum beta-lactamase (TEM-12) and exhibit decreased expression of OmpF. Ceftibuten was unable to select for extended-spectrum beta-lactamase expressing mutants but reduced the expression of two porins, OmpC and OmpF. The stability of ceftibuten to hydrolysis and the difference in the structure of these beta-lactam antibiotics could be responsible for the selection of different mechanisms of resistance.

Ceftibuten stability to active-site serine and metallo-beta-lactamases.

Ceftibuten is an oral third-generation cephalosporin active against a wide range of bacteria and shows an improved stability to hydrolysis by several beta-lactamases because of the carboxyethilidine moiety at position 7 of the ss-acyl side chain. The kinetic interactions between ceftibuten and active-site serine and metallo-ss-lactamases were investigated. The activity of several TEM-derived extended spectrum beta-lactamases (ESbetaLs) against ceftibuten, cefotaxime and ceftazidime was compared using K(m), K(cat) and K(cat)/K(m). Ceftibuten behaved as a poor substrate for class A and B beta-lactamases compared with cefotaxime. The chromosomal class C beta-lactamase from Enterobacter cloacae 908R gave a high K(cat) value (21 s(-1)), whereas there was poor activity with enzymes from Acinetobacter baumannii and Morganella morganii and ceftibuten. Ceftibuten resists hydrolysis in the presence of typical respiratory or urogenital-tract pathogens producing beta-lactamases.

Bactericidal activity of levofloxacin and ciprofloxacin on clinical isolates of different phenotypes of Pseudomonas aeruginosa.

Levofloxacin has been reported to have in vitro activity against both gram-positive and gram-negative bacteria. A recent survey carried out at our Institution showed clinical isolates of Pseudomonas aeruginosa to be more susceptible to levofloxacin than to ciprofloxacin. The in vitro activity of the two fluoroquinolones was evaluated further by looking at their bactericidal activity against two strains of each of the following antibio-phenotypes of P. aeruginosa: levofloxacin- and ciprofloxacin-susceptible, levofloxacin-susceptible/ciprofloxacin-resistant, levofloxacin-susceptible/ciprofloxacin-susceptible and ceftazidime-resistant, (National Committee for Clinical Laboratory Standards susceptibility breakpoints were used). MIC and MBC values were measured and time-kill experiments were carried out. Drugs were used at susceptibility or resistance breakpoint concentrations in the time-kill experiments and results were recorded over 12 h in an attempt to link in vitro results with the clinical situation The polypeptide profiles of outer membrane preparations of the six strains were examined by gel electrophoresis. Levofloxacin was shown to be more bactericidal than ciprofloxacin in the time-kill experiments. No differences were observed between the outer membrane proteins of the six strains. Levofloxacin showed greater bactericidal activity against P. aeruginosa clinical isolates than ciprofloxacin.

Italian survey on comparative levofloxacin susceptibility in 334 clinical isolates of Pseudomonas aeruginosa.

A national survey on susceptibility patterns of 334 Pseudomonas aeruginosa isolates from intensive care units and hematology and oncology wards from 13 Italian hospitals compared the in vitro activity of levofloxacin, an injectable oral fluoroquinolone, to those of ciprofloxacin, ofloxacin, ceftazidime, imipenem, amikacin, and gentamicin. Amikacin and imipenem had the best susceptibility profiles. The activity of levofloxacin was superior to those of the other quinolones and was comparable to that of ceftazidime. The effect of levofloxacin in vitro on P. aeruginosa clinical isolates suggests that further clinical investigations are warranted.

Ceftazidime and aztreonam resistance in Providencia stuartii: characterization of a natural TEM-derived extended-spectrum beta-lactamase, TEM-60.

A plasmid-encoded beta-lactamase produced from a clinical strain of Providencia stuartii has been purified and characterized. The gene coding for the beta-lactamase was cloned and sequenced. It appears to be a new natural TEM-derived enzyme, named TEM-60. Point mutations (Q39K, L51P, E104K, and R164S) are present with respect to the TEM-1 enzyme; the mutation L51P has never been previously reported, with the exception of the chromosomally encoded extended-spectrum beta-lactamase PER-1. Kinetic parameters relative to penicillins, cephalosporins, and monobactams other than mechanism-based inactivators were related to the in vitro susceptibility phenotype.

Interactions of biapenem with active-site serine and metallo-beta-lactamases.

Biapenem, formerly LJC 10,627 or L-627, a carbapenem antibiotic, was studied in its interactions with 12 beta-lactamases belonging to the four molecular classes proposed by R. P. Ambler (Philos. Trans. R. Soc. Lond. Biol. Sci. 289:321-331, 1980). Kinetic parameters were determined. Biapenem was readily inactivated by metallo-beta-lactamases but behaved as a transient inhibitor of the active-site serine enzymes tested, although with different acylation efficiency values. Class A and class D beta-lactamases were unable to confer in vitro resistance toward this carbapenem antibiotic. Surprisingly, the same situation was found in the case of class B enzymes from Aeromonas hydrophila AE036 and Bacillus cereus 5/B/6 when expressed in Escherichia coli strains.

In vitro activity of cefodizime (HR-221) in combination with beta-lactamase inhibitors.

Cefodizime (formerly HR221) was tested either for in vitro microbiological activity or for its stability to beta-lactamases in the presence of two beta-lactamase inhibitors (clavulanic acid, tazobactam). Cefodizime was a poor substrate of class C enzymes but hyperproducer strains were generally resistant with or without a beta-lactamase inhibitor used in combination. On the contrary, class A enzymes were able to hydrolyze cefodizime. However, strains expressing class A beta-lactamase were susceptible to cefodizime in combination with clavulanic acid.

High specificity of cphA-encoded metallo-beta-lactamase from Aeromonas hydrophila AE036 for carbapenems and its contribution to beta-lactam resistance.

The Aeromonas hydrophila AE036 chromosome contains a cphA gene encoding a metallo-beta-lactamase highly active against carbapenem antibiotics. This enzyme was induced in strain AE036 to the same extent by both benzylpenicillin and imipenem. When the cphA gene was inserted into plasmid pACYC184, used to transform Escherichia coli DH5 alpha, the MICs of imipenem, meropenem, and penem HRE664 for recombinant clone DH5 alpha(pAA20R), expressing the Aeromonas metallo-beta-lactamase, were significantly increased, but those of penicillins and cephalosporins were not. When the metallo-beta-lactamase purified from E. coli DH5 alpha(pAA20R) was assayed with several beta-lactam substrates, it hydrolyzed carbapenems but not penicillins or cephalosporins efficiently. These results demonstrate that this metallo-beta-lactamase possesses an unusual spectrum of activity compared with all the other class B enzymes identified so far, being active on penems and carbapenems only. This enzyme may thus contribute to the development of resistance to penems and carbapenems but not other beta-lactams.