Mechanisms of macrolide resistance in clinical pneumococcal isolates in a university hospital, Ankara, Turkey.

Macrolide resistance in Streptococcus pneumoniae is usually caused by the presence of the erm(B) or mef(A) resistance determinants. The aim of the present study was to identify the predominant macrolide resistance mechanisms among erythromycin-resistant S. pneumoniae isolated in a university hospital, Ankara, Turkey. A total of 669 S. pneumoniae strains were isolated from clinical specimens of patients admitted to the hospital between 1994--2002. The minimum inhibitory concentrations (MICs) of penicillin G, erythromycin A and clindamycin were determined by the agar dilution method according to NCCLS guidelines. Ninety-one (13.6%) isolates were resistant to erythromycin. Erythromycin-resistant isolates were examined for their macrolide resistance phenotypes by a triple disc diffusion assay. It assigned 57 (62.6%) of the 91 erythromycin-resistant pneumococci to cMLS(B) phenotype, 19 (20.9%) to iMLS(B) phenotype and 15 (16.5%) to M phenotype. All erythromycin-resistant isolates were analyzed by PCR for the presence of erm(B) and mef(A) determinants. The isolates were characterized for the underlying resistance genotype, with 83.5% having erm(B), 16.5% having the mef(A) genotypes. This study provides further evidence of the dissemination of macrolide-resistant mutants in pneumococci as the use of new, long-acting macrolides increases. This is the first article about MLS(B) resistance phenotypes and genotypes of S. pneumoniae from Turkey and it emphasizes the need for future epidemiological monitoring of macrolide-resistant pneumococci.

In vitro evaluation of faropenem activity against anaerobic bacteria.

Faropenem, a new oral penem with broad spectrum activity, could be used as empirical treatment in infections due to unidentified anaerobes, but only a few investigations have been carried out on these bacteria. The aim of this study was to compare faropenem in vitro activity with that of positive antimicrobial controls (metronidazole, imipenem, meropenem, amoxicillin, amoxicillin-clavulanic acid, ticarcillin-clavulanic acid, cefotetan, cefoxitin and clindamycin) against 462 anaerobic bacterial strains. The reference agar dilution method was used according to the NCCLS standard. Faropenem demonstrated high antimicrobial activity, similar to that of both imipenem and meropenem (faropenem Minimal Inhibitory Concentrations 50% and 90% were 0.12 and 1 mg/L for all Gram-negative anaerobes, 0.25 and 1 mg/L for all Gram-positive anaerobes). Only 5 strains of the Bacteroides fragilis group (1.1% of all anaerobes) were resistant to faropenem, which compared favorably with that of other reference antianaerobic drugs. The results obtained confirm those previously reported.

Affinity of cefmenoxime for beta-lactamases: an analysis.

The interactions of cefmenoxime with beta-lactamases in comparison with cefotaxime, moxalactam, cefoperazone, and ceftazidime have been determined. On-line computerized microacidimetry allowed determination of the affinity of these compounds with the enzymes, which was characterized by Km values. The beta-lactamases that were used were two cephalosporinases and one penicillinase. Within these data, the cephalosporins could be classified into three groups: (1) those with high affinity for the cephalosporinases and very poor affinity for the penicillinase (cefmenoxime, cefotaxime, and moxalactam); (2) those with moderate affinity for the cephalosporinases and very poor affinity for the penicillinase (ceftazidime); (3) those with poor affinity for all enzymes (cefoperazone). In the case of the penicillinase (TEM-1), only cefoperazone was subject to some hydrolysis.

Pharmacokinetic study of cefmenoxime (SCE 1365-CMX) in healthy adults.

Cefmenoxime pharmacokinetics were investigated in six healthy volunteers after intravenous and intramuscular administration of 0.5, 1, and 2 g. Blood and urine samples were analyzed by reversed-phase high-pressure liquid chromatography using ultraviolet detection at 275 nm. The assay is precise and linear up to 200 micrograms/ml-1, with 0.02 micrograms/ml-1 as the limit of detection. Linearity of cefmenoxime kinetics was demonstrated because the area under the plasma concentrations is proportional to studied doses. Eight hours after 1 g of cefmenoxime intramuscularly, mean plasma concentrations are, respectively, 0.6 +/- 0.1 and 0.3 +/- 0.1 microgram/ml-1. Intramuscular cefmenoxime is rapidly absorbed (Ka = 7.28 hours-1) with complete bioavailability (F = 0.99); apparent volume of distribution is 0.35 liters/kg-1 and elimination half-life 1.5 hours. The fraction of cefmenoxime excreted unchanged in the urine after intramuscular administration is 0.72, indicating a major contribution of renal clearance in total clearance. Experimental data after intramuscular administration were well fitted with a two-compartment model.

Classification and structure-activity relationships of fluoroquinolones.

Fluoroquinolones are potent broad spectrum antibacterial agents. Two classifications have been described: chemical and biological. Quinolones can be classified into 4 groups according to their chemical structures: monocyclic, bicyclic, tricyclic and tetracyclic derivatives. Each group can be subdivided into subgroups if a fluorine atom is fixed at the 6-position. The biological classification recognised 4 groups. Groups 1 and 2 are composed of compounds showing limited spectra (Enterobacteriaceae) and groups 3 and 4 contain compounds displaying broad antibacterial spectra. Compounds that are highly metabolised fall into groups 1 and 3 and those poorly metabolised (< 5%) into groups 2 and 4. The structure of fluoroquinolones may help to predict antibacterial activity, pharmacokinetics, physicochemical properties, toxicity and adverse events.

Diffusion of ofloxacin into human lung tissue.

Ofloxacin 200mg twice daily was administered to 17 patients with pulmonary disorders, which necessitated surgery, during the preceding 48 hours and 200mg was administered 1 hour before the operation. During surgery, blood samples and specimens of healthy and diseased lung tissues were taken simultaneously. Ofloxacin levels were determined by HPLC. The mean values of the tissue concentration/plasma concentration ratio were 3.5 +/- 0.4 for the healthy tissue and 3.9 +/- 0.4 for the diseased tissue. These values reflected good penetration of ofloxacin into both healthy and atelectasic pulmonary parenchyma.

Anti-anaerobic activity of antibacterial agents.

The first very effective bactericidal anti-anaerobic drug was metronidazole, introduced in clinical practice in the early 1980s. Sometimes penicillin G and chloramphenicol were used successfully in some anaerobic infections. However, this result was most likely due to Gram-positive anaerobic infections (e.g., Clostridium perfringens). Very rapidly, the anti-anaerobic armamentarium was extended with clindamycin, cefoxitin, imipenem and co-amoxyclav or piperacillin-tazobactam. The resistance rate to metronidazole and imipenem remains low but clindamycin has seen an importance decrease in bacterial susceptibility. New additional drugs could be very helpful to overcome resistance and adverse events. The novelties in this field are fluoroquinolones, which exhibit a good activity against Gram-positive cocci and anaerobes.

Macrolide resistance in Streptococcus pyogenes isolates from throat infections in the region of Aachen, Germany.

Macrolide-resistance was assessed in 216 consecutive Streptococcus pyogenes isolates from throat infections in the region of Aachen, Germany. Seventeen isolates were resistant to erythromycin: 12 isolates revealed a macrolide (M) phenotype and harbored mefA, and five strains expressed an inducible macrolide-lincosamide-streptogramin B (MLSB) phenotype of which four strains harbored ermA(TR) and one strain contained ermB(AM). Telithromycin (HMR 3647) and quinupristin/dalfopristin remained active particularly against the ermA(TR)-containing S. pyogenes isolates studied. Random amplified polymorphic DNA analysis identified multiple clones among erythromycin-resistant strains, but did not discriminate beyond the emm-type. mefA was present in three isolates either with emm2, emm12, or emm75, and in nine isolates with emm4. All four strains with ermA(TR) contained emm77, and the single strain with ermB(AM) harbored emm1. Despite the relative low rate of macrolide-resistance, these data suggest that at least three different macrolide-resistance determinants are prevalent in Germany and that mefA has spread rapidly into multiple clones of S. pyogenes.

Viridans group streptococci: a reservoir of resistant bacteria in oral cavities.

The worldwide spread of erythromycin A-resistant streptococci, including Streptococcus pneumoniae, is of concern. Many studies have demonstrated that the viridans group streptococci can be a reservoir of erythromycin A resistance. Within oral streptoccoci, an important difference in the susceptibility pattern has been noted. The purpose of this short editorial is to highlight the importance of this group of bacteria as a reservoir of resistance to erythromycin A and the possible transfer of resistance to S. pneumoniae and S. pyogenes.

Bacillus anthracis and antibacterial agents.

Anthrax is one of the oldest threats to humankind, and remains endemic in animals in many parts of the world. Human cases are infrequent, and some result from biological warfare. This review summarizes the current knowledge on the antibacterial activity of available antibiotics. For potential use in the most severe cases of anthrax, antibacterials need to exhibit potent in vitro activity, intracellular bioactivity, and suitable locations in lymph nodes. In animal models, it has been shown that doxycycline and fluoroquinolones are the most active compounds. There is a lack of data for animal models for macrolides and ketolides, some of them exhibiting good in vitro activity. However, systemic anthrax (inhalation or gastrointestinal) is mainly due to anthrax toxin, and therapy directed against intoxication is needed as basic treatment.

Ketolides-telithromycin, an example of a new class of antibacterial agents.

Ketolides are new medicinal chemical entities. They are obtained by removing the 3-L-cladinose sugar moiety from erythronolide A and oxidation of the resulting 3-hydroxyl. They were designed to overcome erythromycin A resistance within Gram-positive cocci. The 3-keto group is responsible for the lack of induction of macrolide resistance, high stability in acidic environments, and the ability to overcome resistance due to methylation of 23SrRNA. The C11-C12 carbamate ketolides are able to overcome efflux and hydrolysis mechanisms of resistance and possess additional mechanisms of action at the ribosome level in comparison with erythromycin A. The nature of the side-chain substituting the C11-C12 carbamate residue is responsible for enhancing the in vitro and in vivo activities in comparison with clarithromycin, for the pharmacodynamic and pharmacokinetic properties, for the intracellular features, and for tolerance. This C11-C12 side-chain is supporting the development of new ketolides.

Preparation of stock solutions of macrolide and ketolide compounds for antimicrobial susceptibility tests.

Stock solutions of telithromycin, ABT-773, azithromycin, clarithromycin, erythromycin, roxithromycin and dirithromycin were each prepared with eight different combinations of solvents and diluents. Broth microdilution trays were then prepared and frozen at -60 degrees C. Standard quality control strains were evaluated periodically during a 12-week storage time. There were no significant changes in MICs with different solvents and diluents. It was concluded that the easiest approach was to dissolve each compound in water with a small volume (< 2.5 microL/mL) of glacial acetic acid added in a dropwise fashion, followed by further dilutions in deionised water.

Fluoroquinolones: mechanisms of action and resistance.

The mechanism of action and the mechanism of resistance of the 4-quinolones are complex and poorly understood. The first barrier these molecules must cross is the bacterial outer membrane. In gram-negative species, 4-quinolones pass through either the porins or lipopolysaccharides (or both) depending on their chemical nature. The cellular target is the DNA of the bacterial chromosome. 4-Quinolones mainly modify the activity of topoisomerase II or DNA gyrase, but also that of topoisomerase I and IV. Any damage to the bacterial genome will induce a stress response which consists in the SOS response, chaperonin synthesis or the generation of oxygen free radicals. Peptidoglycan synthesis is also modified, as shown by the inhibition of PBP(3) activity. Mechanism of resistance involves mutations on gyrA and gyrB genes. gyrA Mutations on the chromosome are responsible for a high level of resistance due to a modification of the A subunit of DNA gyrase, mutations on gyrB gene are responsible for a low level of resistance; the combination of both mutations leads to a high level of resistance. Other mutations are responsible for increasing the MIC, such as a norA mutation in S. aureus. The antibacterial activity of the various molecules is different and, as a result, there is not a single mechanism of action or resistance, but rather a common trunk on which additional mechanisms are grafted.

Classification of oral cephalosporins. A matter for debate.

There are many cephalosporins available and various ways of classifying them for clinical use. Oral cephalosporins probably need a classification of their own. This informal discussion was prompted by the appearance of the recommendations of an expert committee of the Paul Ehrlich Gesellschaft. The views of several other commentators are included. There is considerable individual variation in preference for different styles of classification depending on what the classification is for.

Cephems: fifty years of continuous research.

Since 1964, seven waves of parenteral cephems have been reported. All of them were designed to meet medical needs. The first (group I) and the third (group III) waves were very successful and drugs belonging to group III are widely used in the treatment of severe infections. A new series of compounds (group VII), with a new compound underdevelopment was designed for the treatment of Staphylococcus aureus strain resistant to methicillin but also to glyco- and lipoglycopeptides. By modifying the substituent at position 3 and 7 of the cephems rings optimal moieties have been fixed leading to potent anti-Gram-positive drugs. Alterations of substituents are still in progress to obtain optimal anti-Gram-positive (anti-MRSA) compounds. The first oral cephem cephalexin was introduced in clinical practice in 1967. Since this time, many esterified and non-esterified cephems have been synthesized and introduced in clinics. There are two groups of compounds, alpha-amino and non-alpha-amino cephems which are classified in six groups according to their chemical structure. The absorption route was explored and three transporting systems have been described according to the physico-chemical properties of these compounds, in addition prodrugs are passively absorbed.

Antibacterial activity of roxithromycin: a laboratory evaluation.

Roxithromycin, formerly known as RU 28965 (9-[O [(2-methoxyethoxy)methyl]oxime]-erythromycin), is a novel 14 atom-membered semisynthetic macrolide with an antibacterial spectrum directed towards Gram-positive cocci and bacilli, Gram-negative cocci and some Parvobacteriaceae. The in vitro antibacterial activity of roxithromycin was compared with those of erythromycin and spiramycin against 275 clinical isolates by using 2-fold broth macro-dilution tests. The antibacterial spectrum of roxithromycin and erythromycin were qualitatively comparable (including the bacteriostatic type of activity and the profile of resistance), but minimal inhibitory concentrations of erythromycin were generally one half those of roxithromycin, except for Corynebacterium sp. and Bacteroides fragilis against which the new macrolide was more active. On the other hand, roxithromycin exhibited a superior in vivo antibacterial activity in laboratory animals, being up to six times more potent than erythromycin in curing experimentally infected mice. Roxithromycin showed high blood levels and long half-lives of elimination in rodents after oral administration, and its bioavailability amounted to 72% in mice and 85% in rats, compared to less than 10% for erythromycin. Roxithromycin was widely distributed throughout the body with a high degree of penetration into all tissues, particularly in the lungs.

Behaviour of cefmenoxime towards beta-lactamases.

Using several well-characterized beta-lactamases isolated from Gram-negative bacteria, the interactions of cefmenoxime, a new methoxy-imino-amino-2-thiazol cephalosporin were compared with those of cefotaxime, lamoxactam, cefoperazone and ceftazidime. On-line computerized microacidimetry allowed determination of the affinity of these compounds for the enzymes, which was characterized by Ki values. Microacidimetry showed poor interactions of cefmenoxime with penicillinase TEM-1 (low Vm, poor affinity) whereas it showed a high affinity for the cephalosporinases, as is also the case for cefotaxime or lamoxactam. Both cefmenoxime and cefotaxime showed relative susceptibility in Masuda's double disc technique.

[Role of Clostridium and its toxin in pseudo-membranous colitis (author's transl)]. / Rôle de Clostridium difficile et de sa toxine dans les colites pseudo-membraneuses.

At present many authors consider that pseudo-membranous colitis is of bacterial origin. The main pathogenic agent is Clostridium difficile. It is not easy to isolate this organism in the stool, selective media are under study. It liberates a lipo-glycoprotein exotoxin during lysis. It is only partially purified, its structure is not fully elucidated. Its molecular weight is not yet precisely determined. It consists of several polymerised polypeptide fragments of molecular weight 50 000. It is a thermolabile acid and alkaline sensitive cytotoxin which acts on the cell membranes and the ileo-caeco-colonic mucosa of man and animals. Clostridium difficile is transmissible by a small number of high risk carrier subjects who are potentially patients with pseudo-membranous colitis. Antibiotic therapy may lead to unbalance of the ecosystem represented by the bacterial flora of the digestive tract and favour the multiplication of a resistant strain to the administered antibiotic. The appearance of pseudo-membranous colitis requires the association of sufficient bacterial development (equal or greater than 10(7) germs per gram of stools) and the liberation of a cytotoxin. The pathogenic treatment consists of antibiotic therapy by Vancomycin or Metronidazole which seems, at present, the most active on the germs and a toxin absorbent, such as Cholestyramine, Coliptol hydrochloride or Heavy metals.

[Effect of rheumatoid factors on results for determination of antitoxoplasms IgM antibodies by immunofluorescence and agglutination technics (author's transl)]. / Influence des facteurs rhumatoïdes sur la détermination des anticorps antitoxoplasmes IgM par immunofluorescence et par agglutination.

Simultaneous presence of antitoxoplasm IgG antibodies and of rheumatoid factors gives rise to an IgM-type immunofluorescence reaction in 44 p. cent of cases. Three experiments showed that association of the two antibodies can give rise to false positives in the Remington test ; the mixture of the two antibodies renders the reaction positive in the majority of cases. On the other hand, after absorption of negative rheumatoid factors and separation of serum IgM and IgG, IgM fluorescence is no longer seen ; if they are again mixed, then a positive reaction will result. Interference of antitoxoplasm IgG antibodies and rheumatoid factors does not occur in direct toxoplasm agglutination tests. The presence of the rheumatoid factor is rare in the pregnant female. Nevertheless, it is necessary to test for rheumatoid factors using the latex test. If results are here positive, then serum should be absorbed on aggregated human IgG and the Remington test repeated. The probably rare simultaneous presence of antitoxoplasm IgG antibodies and of rheumatoid factors cannot be differentiated from these false positives without fractionation of the serum, separating IgM and IgG.

[Antibiotherapy in the past, present and future]. / L'Antibiothérapie au passé, au présent et au futur.

The antibiotic era started in 1941. Several antibiotics were obtained from the fermentation broth, either from moulds (penicillin) or from Actinomyces species (erythromycin). Numerous antibacterial agents were prepared synthetically such as benzylpyrimidines, 4-quinolones, nitroheterocyclic derivatives, have been developed. Consecutive "waves" of newer antibiotics was helpful to solve some infectious problems but simultaneous emergence of resistant strains has occurred amongst initially susceptible bacteria. This imposes reconsideration of our therapeutic armamentarium and a search for new compounds as well as for new concepts in anti-infective therapy.