Resistencia a los β -lactámicos en enterococos / Resistance to β-lactams in enterococci

Enterococci are intrinsically resistant to several antimicrobial classes and show a great ability to acquire new mechanisms of resistance. Resistance to β-lactam antibiotics is a major concern because these drugs either alone or in combination are commonly used for the treatment of enterococcal infections. Ampicillin resistance, which is rare in Enterococcus faecium occurs in most of the hospital-associated Enterococcus faecium isolates. High-level resistance to ampicillin in E. faecium is mainly due to the enhanced production of PBP5 and/or by polymorphisms in the beta subunit of this protein. The dissemination of high-level ampicillin resistance can be the result of both clonal spread of strains with mutated pbp5 genes and resistance horizontal gene transfer.

Los enterococos son intrínsecamente resistentes a varias clases de antimicrobianos y presentan una gran capacidad para adquirir mecanismos de resistencia. La resistencia a los antibióticos p-lactámicos es preocupante porque estos fármacos solos o combinados se usan comúnmente para el tratamiento de las infecciones enterocócicas. La mayoría de los aislamientos hospitalarios de Enterococcus faecium presentan resistencia a la ampicilina, la cual es rara en Enterococcus faecalis. El alto nivel de resistencia a la ampicilina en E. faecium se debe principalmente a la hiperproducción de PBP5 y/o a polimorfismos en la subunidad beta de esta proteína. La propagación de esta resistencia puede deberse tanto a la diseminación clonal de cepas con genes pbp5 mutados como a la transferencia horizontal de genes.

Simple genetic selection protocol for isolation of overexpressed genes that enhance accumulation of membrane-integrated human G protein-coupled receptors in Escherichia coli.

The efficient production of membrane proteins in bacteria remains a major challenge. In this work, we sought to identify overexpressed genes that enhance the yields of recombinant membrane proteins in Escherichia coli. We developed a genetic selection system for bacterial membrane protein production, consisting of membrane protein fusions with the enzyme beta-lactamase and facile selection of high-production strains on ampicillin-containing media. This system was used to screen the ASKA library, an ordered library of plasmids encoding all the known E. coli open reading frames (ORFs), and several clones with the ability to accumulate enhanced amounts of recombinant membrane proteins were selected. Notably, coexpression of ybaB, a gene encoding a putative DNA-binding protein of unknown function, was found to enhance the accumulation of a variety of membrane-integrated human G protein-coupled receptors and other integral membrane proteins in E. coli by up to 10-fold. The results of this study highlight the power of genetic approaches for identifying factors that impact membrane protein biogenesis and for generating engineered microbial hosts for membrane protein production.

Interaction between 3,5-diacetyl-1,4-dihydropyridines and ampicillin, and erythromycin on different E. coli strains.

Eleven analogues of nifedipine (NP) showed synergistic interactions with ampicillin (Ap) and erythromycin (Er) on Escherichia coli K12LE140/F'lac. The antibacterial effect of Ap was enhanced by most analogues but compound (G9) and (+/-)-verapamil (VP) were antagonistic. Two of the 11 compounds (G7, G8) were synergistic with Er and four were additive. With a sensitive clinical isolate of E. coli Gy-1/Ap(sens)Er(res), compound G1 antagonized the antibacterial effect of Ap and a synergistic effect was found in the combination of Er with G4, G5, G6 or G7. None of the drugs had any effect on a multidrug resistant (MDR) clinical isolate of E. coli Gy-2/Ap(res)Er(res).

Inhibition of peptidoglycan hydrolase activity in vivo and in vitro by energy uncouplers in Escherichia coli.

The effects of energy uncouplers on in vivo and in vitro peptidoglycan hydrolase activities in Escherichia coli were determined. Sodium azide, potassium cyanide, and carbonyl cyanide m-chlorophenylhydrazone all inhibited ampicillin-induced lysis of exponential phase cultures, even when they were added to lysis-committed cultures. These energy uncouplers also inhibited the solubilization of radiolabeled peptidoglycan by bacterial suspensions that had been treated with 5% trichloroacetic acid by the method of Hartmann et al.3 to activate the peptidoglycan hydrolases. Therefore, the in vivo and in vitro activities of peptidoglycan hydrolases in E. coli are dependent on membrane energization.

Modulation of antimicrobial effects of beta-lactams by amino acids in vitro.

Glycine as well as 11 and 10, respectively, out of a total of 12 D-amino-acids tested increased the antimicrobial efficacy of imipenem (IMI) and of ampicillin (AMP) using the serosensitive strain E. coli ATCC 8739. D-proline was ineffective in assays with IMI as well as D-proline and D-leucine in assays with AMP. - In contrast, L-amino-acids behaved differently: In assays with IMI, 9 out of 13 isomers were ineffective whereas 3 were antagonistic (L-phenylalanine, L-serine, L-tryptophan). In combination with AMP, however, 10 L-amino acids had an antagonistic effect and 2 (L-leucine, L-methionine) were ineffective. L-alanine was an exception and showed a synergism with both antibiotics which was assumed to have been due to a racemase activity of cells. - Seroresistance of E. coli apparently reduced the synergistic effect of glycine and beta-lactams. - Glycine, alanine and tryptophan lost their typical synergistic or antagonistic effect with AMP when tested as di- or tri-amino-acid compounds. This was not the case with di-L-alanine - It is supposed that the synergistic effect of glycine or of D-amino-acids with beta-lactams can be explained mainly by an inhibition of carboxypeptidases.

The activity of vancomycin and teicoplanin alone and in combination with gentamicin or ampicillin against Streptococcus faecalis.

The in vitro activities of vancomycin and teicoplanin against 56 strains of Streptococcus faecalis were compared. Killing curves showed that vancomycin and teicoplanin were bacteriostatic and that synergy was achieved when each was combined with gentamicin. The bactericidal activity displayed by 4 mg/l of ampicillin against Streptococcus faecalis was antagonized by 4 mg/l of vancomycin and 0.5 mg/l of teicoplanin respectively.

Comparative pharmacokinetics of dicloxacillin and ampicillin between individual and combined doses.

Pharmacokinetics of dicloxacillin and ampicillin dosed individually and in combination were investigated by means of moment analyses of urinary excretions of intact forms and metabolic products of parent penicillins. Comparison of excretion profiles between individual and combined doses to human subjects indicated that the transformation of ampicillin to penicilloic acid and subsequent conversion to secondary metabolite are suppressed by the simultaneous dose of dicloxacillin, while the total excretion ratio to dose and the mean residence times in the body remain almost unchanged. The excretion profiles of dicloxacillin and metabolites are not significantly affected by the combined dose.

Hydroxylamine inactivation of cephalosporins: nucleophilic attack on beta-lactam structures.

Cephalosporins are not degraded by hydroxylamine (NH2OH) in neutral and acidic solutions. Their reaction with NH2OH in slightly alkaline solutions leads to microbiological inactivation which seems to be a structure dependent phenomenon. In these experiments the mandelic acid-type compounds appear to be quite stable to the effect of NH2OH, whereas, cefazolin is gradually degraded and the straight chain-containing cephalosporins are variably inactivated. The phenylglycine-type oral cephalosporins were generally sensitive to the alkaline conditions used in these tests and apparently are not inactivated by NH2OH. On the contrary, the phenylglycine-type cephalosporins seem to be somewhat stabilized in the presence of NH2OH.

[Rapid methods of detecting enzymatic resistance to ampicillin and chloramphenicol in Haemophilus influenzae]. / Méthodes rapides de détection de la résistance enzymatique a l'ampicilline et au chloramphénicol chez Haemophilus influenzae.

Several methods used for the detection of beta-lactamase activity in Haemophilus influenzae are described. The rapid iodemetric, acidimetric, and chromogenic cephalosporin techniques are specific tests for the presence of beta-lactamase. The Gots test can also be used for the detection of enzymatic resistance to ampicillin and chloramphenicol.

[Migration of the ampicillin transposon in enteropathogenic Escherichia]. / Migratsiia ampitsillinovogo transpozona u énteropatogennykh ésherikhii.

Migration of Tn 1 from plasmid RP4 into the chromosome of enteropathogenic Escherichia (EPE) of serogroups O124 and O111 was studied. E. coli K 12 LC 411 (RP4) was used as the donor. It was shown that the transposition rate markedly differed depending on the period of ;the cell isolation from Tn 1 in the chromosome, i. e. during conjugation or after several subcultures of the transconjugants from the autonomic plasmid onto the selective media. During the conjugation process the migration rate of Tn 1 was equal to 0.5 and 0.8 per cell acquiring the plasmid. The transposition rate in the EPE carrying the R factor for a long period of time was 2.4 . 10(-2) and 1.4 . 10(-2) respectively for each serogroup. The above differences in the migration rate of Tn 1 were not concerned with changes in the environment.

In vitro antagonism by erythromycin of the bactericidal action of antimicrobial agents against common respiratory pathogens.

Ten strains each of Staphylococcus aureus, Haemophilus influenzae, Enterobacter spp., Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, and Streptococcus pneumoniae were tested in vitro against erythromycin combined with ampicillin, cefamandole, or gentamicin. Antagonism by erythromycin occurred with 47% of the combinations involving strains of S. aureus and to a lesser degree with H. influenzae. Synergy occurred most commonly with H. influenzae (27%). The high frequency of antagonism and synergy with these organisms was associated with a broad range of bacteriostatic action by erythromycin against these same bacteria. The implications for the treatment of pneumonia are discussed.

[Auxotrophic mutations formed on the incorporation into the E. coli C600 chromosome of transposon Tnl that determines ampicillin resistance]. / Mutatsii auksotrofnosti, voznikaiushchie pri vstraivanii v khromosomu E. coli C600 transpozona Tnl, opredeliaiushchego ustoichivost' k ampitsillinu.

Incorporation of transposone Tnl of ampicillin resistance into the chromosome of E. coli C600 resulted in formation of at least 8 types of auxotrophic mutants. The transposone incorporates mainly into the loci the damage of which induces proline deficit (20 per cent). The sites of incorporation of transposone Tnl in the chromosomes of E. coli C600 and E. coli JC411 did not coincide.

Response of Escherichia coli to Beta-lactam antibiotics: effect of the concomitant presence of staphylococci exhibiting inducible Beta-lactamase activity.

The response to Beta-lactam antibiotics of mixed cultures of Escherichia coli and Staphylococcus epidermidis was studied in static culture and in an in vitro model which simulates the dynamic conditions in which bacteria are exposed to antibiotics in the treatment of urinary infection. In static cultures, the concomitant presence of staphylococci exhibiting inducible Beta-lactamase activity substantially reduced the efficacy of benzylpenicillin and ampicillin (but not cefuroxime) against E. coli. In the conditions of the bladder model some interference with the activity of Beta-lactam antibiotics by Beta-lactamase producing staphylococci was also demonstrated. Nevertheless, relatively modest doses of ampicillin were still able to suppress growth of susceptible E. coli for periods exceeding the normal interdose interval, even in the presence of enzyme-producing staphylococci.

[RP4 factor integration with E. coli chromosome]. / Integratsiia faktora RP4 s khromosomoi E. coli.

Integration of R-factor RP4 with the chromosome of E. coli was studied with the use of replication thermosensitive mutant pEG1 of this factor. It was found that the frequency of integration of factor pEG1 containing the ampicillin transposone Tn1 with the chromosome of bacteria JC411 carrying transposone Tn1 previously inserted into it was very high and markedly exceeded that of its insertion into the same chromosome but not carrying this transposone. The frequency of factor pEG1 insertion into the chromosome of bacteria JC 1553 rec A defective with respect to genetic recombination was less than 2.10(-5) and did not depend on the presence of transposone Tn1 in it. Probably, insertion of factor RP4 into the bacterial chromosome may be realized through the rec A-dependent process of recombination between transposone Tn1 previously translocated into the chromosome and the same transposone contained in R-factor.

Possible beta-lactamase activities detectable in infective clinical specimens.

Biological beta-lactam antibiotic-inactivating activities were detected in bacteriuria and suppurating pleural fluids. Clinical specimens were sterilized with membrane filters and the amounts of ampicillin and/or cephalothin which were being inactivated by 1 ml of each filtrate were determined. In general, filtrates which originally yielded Klebsiella sp. tended to show activity against ampicillin; whereas those yielding Enterobacter sp. and Pseudomonas aeruginosa showed activity against cephalothin.

[Deacylating activity of sera as a cause for the ineffectiveness of the treatment of experimental plaque in white mice with benzylpenicillin]. / Deatsiliruiushchaia aktivnost' syvorotok kak prichina neéffektiv-nosti lecheniia éksperimental'noi chumy belykh myshei benzilpenitsillinom

It was shown that serum of albino mice infected with plague microbe cells inactivated benzylpenicillin. Such deacetylating activity reached its peak by the 3rd day and after that decreased reaching by the 5th--7th day the level registered in non-infected animal and being apparently of non-specific character. Ampicillin proved to be 2 times more resistant to the effect of serum acylase as compared to benzylpenicillin. It was supposed that the ability of serum of infected animals to inactivate benzylpenicillin by splitting off phenylace acid was the cause of ineffective treatment of experimental plague of albino mice with comparatively low doses of the drug.