Four years of monitoring antibiotic resistance in microorganisms from bacteremic patients.

From the second semester of 2002 to the end of the first semester of 2005, a total of 2544 bacterial strains were isolated from the blood stream of patients with clinical sepsis and bacteremia hospitalized in six University Hospitals in the Slovak Republic. Almost 30% of strains were coagulase-negative staphylococci (CONS), about 14% were Staphylococcus aureus and, of the Gram-negative bacteria, up to 9% were Klebsiella pneumoniae. All CONS, S. aureus and Enterococcus spp. strains were found to be still susceptible to vancomycin, but the resistance of CONS and/or S. aureus to macrolides and fluoroquinolones dramatically increased during the period of this study. Among Gram-negative bacteria, increasing levels of resistance to higher generation cephalosporins, to fluoroquinolones resistance in Pseudomonas aeruginosa and Acinetobacter spp. to meropenem was recorded, which is alarming. The results were periodically submitted to cooperating hospitals with proposals for rationalizing the prophylactic and general use of indicated antibiotics as well as for improving hospital hygiene measures and anti-epidemic practices.

Monitoring of antibiotic resistance in bacterial isolates from bacteremic patients.

The aim of the study was to monitor the prevalence of pathogens and development of resistance in bacteria isolated from bacteremic patients. Five University Clinics and/or Regional Hospitals in the Slovak Republic participated in the study and a total of 421 isolates were collected in the second half of the year 2002. The most prevalent organisms were coagulase-negative staphylococci (CONS) (19%), Staphylococcus aureus (18.3%), among Gram-negative bacteria Escherichia coli (13.3%), Klebsiella pneumoniae (11.4%) and Pseudomonas aeruginosa (7.8%) followed by enterococci, Acinetobacter baumannii and Enterobacter sp. All CONS and S. aureus were susceptible to vancomycin; resistance to oxacillin was observed for 55% of the CONS and only for 4% of S. aureus isolates. A higher prevalence of resistance to erythromycin, clindamycin, gentamicin and ofloxacin was found in CONS in comparison to S. aureus. Enterococcus sp. isolates were fully susceptible to vancomycin and teicoplanin. Gentamicin, amoxicillin/clavulanate, third generation cephalosporins and ciprofloxacin showed good activity against E. coli. Although 17% of K. pneumoniae isolates were resistant to ciprofloxacin, it was the most effective drug against K. pneumoniae; the prevalence of resistance to other antibiotics was rather higher. Gentamicin and ciprofloxacin were the most active against Enterobacter sp. isolates and ceftazidime and meropenem against P. aeruginosa.

First occurrence of transferable extended-spectrum beta-lactamase hydrolyzing cefoperazone in multiresistant nosocomial strains of Klebsiella pneumoniae from two hospitals in Czech and Slovak Republics.

The occurrence of positive synergy between antibiotic discs of amoxicillin/clavulanate and cefoperazone was registered in two Klebsiella pneumoniae strains, isolated from hospitals in Czech and Slovak Republic, indicating the presence of genes coding for an extended-spectrum beta-lactamase active also against cefoperazone, a broad-spectrum cephalosporin. Sulbactam inhibited the hydrolysis of cefoperazone by cell-free lysates of these strains which substantiates its use in combination with cefoperazone. Resistance to cephalothin, cefotaxime, ceftazidime, cefoperazone, cefepime and aztreonam was transferred from K. pneumoniae isolates to Escherichia coli K-12 3110 and to Proteus mirabilis P-38 recipient strains.

[New aspects of antibiotic resistance and possibilities of its prevention]. / Novsie aspekty rezistencie na antibiotiká a niektoré moznosti jej limitácie.

New phenomena of the antibiotic resistance in bacteria have recently appeared. The may hold present explosive development of resistance and prevent its transferability from multiple drug resistant bacteria to still sensitive ones. They may prevent the production of so-called extended-spectrum beta-lactamases (ESBLs) among Enterobacteriaceae producing resistance virtually to all penicillins and cephalosporins with exception of those antibiotics potentiated by clavulanic acid or sulbactam, the resistance to vancomycin in enterococci and staphylococce, and the resistance of Stenotrophomonas maltophilia. Factors participating on the development of resistance include: a) transferability of resistance genes among bacteria which explosively change susceptible strains to resistant ones, b) dosage and types of antibiotics which cause the selection pressure to certain species of bacteria, c) level of organization and strict adherence to hygienic and anti-epidemic regimen starting with the entry of patients into the hospital. Analyses are necessary to check whether the patient brings resistant bacteria with a transferable resistance (with ESBLs) into the hospital. Preventive measures would be strictly applied to stop the clonal spread of resistant strains among the patients and/or hospital environment, which occurs if these strains have such opportunity. Last, but not least to be considered is the dosage, composition and rationality of administration of antibacterials, mainly in post-operative prophylaxis in intensive care units, in so-called empirical usage, etc. At the same time, it would be highly unethical to hesitate with application of antibacterials to patients when it is justified, necessary and rational. Hospital antibiotics policy should rationally decide between these alternatives in each application of antibiotics or their combinations.

Transferable antibiotic resistance in multiresistant nosocomial Acinetobacter baumannii strains from seven clinics in the Slovak and Czech Republics.

Sixty-seven multiresistant nosocomial Acinetobacter baumannii isolates from patients hospitalized mostly in intensive care units of seven clinics in Slovak and Czech Republic were tested to determine their ability to transfer antibiotic resistance. All isolates were resistant to kanamycin, ticarcillin, cephalothin, cefotaxime, ceftazidime, aztreonam and susceptible to carbapenems, sulbactam and ampicillin/sulbactam. Sixty-five out of 67 strains transferred resistance determinants to Escherichia coli K-12 and Proteus mirabilis P-38 recipients. Analysis of selected transconjugants by an indirect selection method showed a more variable pattern of transferred resistance determinants. The clonal spread of strains transferring resistance seems to be an additional risk for occurrence of strains resistant to ceftazidime and aztreonam.

Sudden increase in Pseudomonas aeruginosa nosocomial strains with broad host range transfer of antibiotic resistance.

We investigated transfer of antibiotic resistance from 51 multiply resistant strains of Pseudomonas aeruginosa isolated from seriously ill patients in the Frankfurt University Clinics. Nine isolates directly transferred resistance to three recipient strains used. Ticarcillin and cephalothin resistance determinants were accepted from eight isolates, and in one case a kanamycin resistance determinant was transferred. The total spectrum of resistance transferred demonstrated that several donor strains transferred a different set of resistance determinants to all three recipient strains. Two P. aeruginosa isolates transferred spectrum of seven resistance determinants including ceftazidime, cefepime and aztreonam, three isolates transferred five determinants and four isolates transferred four resistance determinants. The fact that identical spectra of multiple drug resistance were transferred to recipient strains belonging to three different species (Escherichia coli, Proteus mirabilis and P. aeruginosa), indicates a broad host range in all three transferable genetic elements not observed in previous transfers from P. aeruginosa strains.

Host range variability in the transfer of antibiotic resistance determinants from Pseudomonas aeruginosa strains.

Twelve multiply drug-resistant strains of Pseudomonas aeruginosa, isolated in Frankfurt University Clinics during the year 1998, transferred determinants of antibiotic resistance with remarkable variability in the host range of transferred plasmids. As a new phenomenon, two of them transferred resistance genes to both groups of recipients, i.e. to Enterobacteriaceae as well as to a strain of P. aeruginosa. Six strains had more restrictive activity and transferred resistance determinants to Enterobacteriaceae only, while four additional strains demonstrated transferability only to P. aeruginosa. Two isolates transferred imipenem resistance and one cefepime resistance. Differences in selection pressure occurring during therapy of individual patients seems to be one of the possible factors influencing the spectrum of antibiotic resistance genes present in P. aeruginosa and their transfer to susceptible strains.

Low-Frequency transduction of imipenem resistance and high-frequency transduction of ceftazidime and aztreonam resistance by the bacteriophage AP-151 isolated from a Pseudomonas aeruginosa strain.

Bacteriophage AP-151, isolated from a multidrug resistant Pseudomonas aeruginosa strain, was found to transduce antibiotic resistance determinants to recipient strains of P. aeruginosa. Resistance to cefotaxime, ceftazidime, aztreonam, imipenem and meropenem was transduced as a block, at different frequencies, to two P. aeruginosa strains. Resistance was two logarithms higher (in the range 10(-5)) for cefotaxime, ceftazidime or aztreonam than for imipenem in recipient strain PAO-1670. The frequency of transduced imipenem resistance was also lower in recipient strain ML-1008. This phenomenon reflects the difference in the lytic activity of AP-151 in both strains, as the titer of the AP-151 phage in the PAO strain was found to be restricted to 10(-4)-10(-5) in contrast to the titer of the same phage in the ML strain which was 10(-10). The limited lytic activity in the PAO recipient strain was correlated with higher transducing activity. It can be concluded that some wild-type bacteriophages of P. aeruginosa might have highly individual relations between lytic and transducing activity in various potential recipient nosocomial strains of P. aeruginosa. The nature of resistance to ceftazidime and imipenem was studied using clavulanate and EDTA as inhibitors of individual class of beta-lactamases, indicating the presence of extended-spectrum beta-lactamase and a metallo-beta-lactamase in this isolate.

[60 years' of antibiotics--resistance: a time bomb for the third millenium]. / Sestdesiat rokov antibiotík--rezistencia: casovaná bomba pre tretie tisícrocie.

More then 50 years of antibiotic era has brought to clinical use many antibiotics with, specific and also with broad spectrum activity. Excessive use of antibiotics brought and brings serious problems--the resistance of many important bacteria and so the loss of effectivity of these scarce substances. Significant negative role in this process has the transferability of the resistance genes between bacteria. Bacterial strains are able to accept block of genes for multiresistance, exchange them inter species and spread to susceptible ones. There is an important task for the 3rd millennium to reduce the prevalence of the antibiotic resistance. Changes of the empiric treatment regiments (reduction in antibiotic use, antibiotic rotation, infection control, monitoring of resistance) and the strict hospital hygiene could help to solve the problems of antibiotic resistance in the future.

[Reservoirs, interactions and stability of genetic resistance to antibiotics. The "easy to get--hard to lose" syndrome]. / Rezervoáry, interakcie a stabilita génov rezistencie na antibiotiká. Syndróm "easy to get--hard to lose".

Enthusiasm after discovery of antibiotics and their use in clinical practice led to presumption that problems of bacterial infections will be soon resolved and forgotten and attention will be turned to other serious problems, such as viral infections or neoplastic diseases. However, instead of disappearance of bacterial infections, bacterial pathogens become more resistant to many antibiotics. The ability of bacterial strains to acquire resistance genes from other bacteria, even of different species, causes increasing stability of resistance of bacteria. Transferable elements--resistance genes--often interact and create changed structures; this enables to preserve, stabilize, or under special conditions, transfer resistance genes. Transferable elements include plasmids, transposons, integrins and gene cassettes. Conjugation of bacteria, transduction by bacteriophages and transformation are the mechanisms by which these elements are transferred. A very significant property of transferable, mobilizable and transposable genetic systems of resistance is their stability and ability to adapt to new hosts. They do not lose it in the absence of antibiotics. The generally pessimistic view on future antibacterial chemotherapy should be a challenge to prevent the existence and spread of resistant strains of bacteria. It is much simpler and more convenient than "quench the fire" later. Best scheme is to stop resistance before it starts.

[Antibiotic resistance]. / Niektoré problémy rezistencie na antibiotiká.

The authors present a short review of the origin and evolution of antibiotic resistance from the beginning of antibiotic use. Transferable resistance, i.e. spread of resistance genes by mechanisms of the transduction, conjugation or transposition, plays an important role in the process of the development of resistance in susceptible bacterial strains. Equally, chromosomally coded resistance is recently becoming relevant. Large selective pressure of the antibiotics lends mutations of genes coding antibiotic resistance. So, bacterial strains produce a large amount of enzymes, which destroy antibiotic or lose the power for penetration of the antibiotics become completely resistant also to new antibiotics.

Extended-spectrum beta-lactamase producing strains of Enterobacter cloacae transferring resistance to cefotaxime and ceftazidime.

Transferability of ceftazidime resistance and of extended-spectrum beta-lactamase (ESBL) production was recorded in three strains of Enterobacter cloacae from Frankfurt University Clinics. All three strains were resistant to rifampicin, carbenicillin, cephaloridine, cefotaxime, ceftazidime, aztreonam, imipenem and meropenem. They transferred directly determinants of resistance to carbenicillin and cephaloridine to Escherichia coli K-12 No. 185 nal+ recipient strain. All transconjugant colonies also contained determinants of resistance to cefotaxime, ceftazidime and aztreonam. In the second cycle of transfers, determinants of resistance to carbenicillin, cephaloridine, cefotaxime, ceftazidime and aztreonam were transferred en bloc to the recipient Escherichia coli K-12 No. 3110 rif+ indicating that the "transfer factor" (tra+) mediating the process of conjugational transfer had also been co-transferred. Double-disk diffusion test showed the same pattern of production of ESBLs both in original strains and in transconjugants. Transfer of cefotaxime, ceftazidime and aztreonam was accompanied by the transfer of ESBL production.

[Transfer of resistance to 3rd generation cephalosporins and aztreonam in strains of Klebsiella pneumoniae producing extended spectrum beta-lactamases]. / Prenosná rezistencia na cefalosporíny 3. generácie a aztreonam u kmenov Klebsiella pneumoniae produkujúcich beta-laktamázy s rozsíreným spektrom úcinku (ESBL).

The authors demonstrated the transferability of antibiotic resistance genes in nosocomial strains of Klebsiella pneumoniae, isolated from newborn children at the Paediatric University Hospital in Bratislava. Strains were resistant to cefotaxime, ceftazidime and aztreonam. The determinants of resistance (carbenicillin, cephaloridine, cefotaxime, ceftazidime and aztreonam) were transferred to recipient strains of Escherichia coli K-12 and Proteus mirabilis P-38. The transfer of resistance determinant from donor strains was demonstrated by the analysis of the resistance spectrum in transconjugant clones of recipient strains by the method of indirect selection. The ability of production of extended spectrum beta-lactamases (ESBL) was demonstrated by the double disc diffusion test. Synergy between clavulanate and cefotaxime, clavulanate and ceftazidime and/or clavulanate and aztreonam indicated production of ESBL by these strains.

Transduction of antibiotic resistance in Pseudomomas aeruginosa: relationship between lytic and transducing activity of phage isolate AP-423.

Isolation and propagation of a wild type phage, isolate AP-423, from an apparently lysogenic strain of Pseudomonas aeruginosa, resistant to a series of anti-pseudomonadal antibiotics, and its use for transduction of resistance determinants is described. The phage isolate AP-423 showed a phenomenon of host restriction, i.e. it was lysogenic only for some of the recipient strains tested. Its transduction capacity, both in sets of genes transduced and frequency of transduction, was different in two recipient strains of P. aeruginosa. This phage showed also some restriction in titers, to which it could be propagated, only in certain recipient strains.