Comparative in vitro efficacies of various antipseudomonal antibiotics based catheter lock solutions on eradication of Pseudomonas aeruginosa biofilms

Antibiotic lock technique (ALT) may be an adjunct therapy in treating catheter-related infections. The aim of this study was to determine the in vitro stability and efficacy of colistin, meropenem and levofloxacin alone or in combination with clarithromycin or heparin lock solutions against biofilm embedded Pseudomonas aeruginosa strains. The efficacy of antibiotic lock solutions was tested in an in vitro catheter biofilm model against P. aeruginosa isolated from catheter-related bacteremia. We observed that the use of meropenem, levofloxacin or colistin as a lock solution had potent bactericidal effects and could be prevented bacterial regrowth at 96 or 72 hours, respectively. When the tested antibiotics were used in combination with clarithromycin, the combinations were significantly more effective and rapid in eliminating P. aeruginosa colonization in biofilm than each of the antibiotics were used alone. Moreover, tested antibiotics in combination with heparin were not significantly different for killing effect against PA-1 and PA-27853 compared with that of each antibiotics alone. Tested catheter lock solutions may have promising adjuvants for treating infections caused by P. aeruginosa.     

Effect of Temperature on Antimicrobial Susceptibilities of Pseudomonas Species Isolated from Drinking Water

Summary

The susceptibility of 20 strains of Pseudomonas species isolated from drinking waters (4 P. aeruginosa, 7 P. fluorescens, 5 P. stutzeri, 1 P. maltophilia, 1 P. cepacia, 1 P. putida and 1 P. pickettii) to a variety of antibiotics (gentamicin, amikacin, azlocillin, cefotaxime, chloramphenicol and polymyxin B) were determined by Stoke’s method at 20°C, 30°C, 37°C and 42°C.

Minimum inhibitory concentrations (MIC) were determined for aminoglycosides on Mueller-Hinton agar at the above temperatures. There was a significant difference in susceptibility between 20°C or 30°C (most resistant), 37°C (more susceptible) and 42°C (most susceptible) to gentamicin and to a lesser degree to amikacin for P. maltophilia, P. cepacia and most strains of P. fluorescens. The P. aeruginosa, P. stutzeri, P. putida and P. pickettii strains showed no difference in susceptibility at 20°C, 30°C, 37°C and 42°C. The need for standardized conditions with special regard to temperature when antibiotic susceptibility tests are performed for P. maitophilia, P. cepacia and P. fluorescens strains is stressed.     

Effect of subminimal inhibitory concentrations of azithromycin on adherence of Pseudomonas aeruginosa to polystyrene

Pseudomonas aeruginosa forms a bacterial biofilm by producing alginate when it adheres to mucosa or various medical devices. In this study, the effect of subminimal inhibitory concentrations (subMICs) of azithromycin (AZM) on the biofilm formation and in vitro adherence to polystyrene of 14 wild-type P. aeruginosa strains was studied. A total of 35 P. aeruginosa isolates from clinical specimens were used. Glycocalyx production was determined by the tube method, and bacterial adherence to the wells of flat bottom polystyrene tissue culture plates was estimated by the spectrophotometric method. Compared to the control, the adherence ability to polystyrene was inhibited by incubation with subMICs of AZM in a dose-dependent manner. These results support the findings of other investigators suggesting that AZM in subinhibitory concentrations may be useful in the prevention or treatment of biofilm-associated infections due to P. aeruginosa.     

Relationship of antibiotic resistance phenotype to the R-pyocin susceptibility pattern in clinical isolates of Pseudomonas aeruginosa

One hundred sixteen clinical isolates of Pseudomonas aeruginosa were collected from 7 hospitals in Athens. All strains were studied for their susceptibility to cefotaxime, ceftazidime, carbenicillin, aztreonam, imipenem, nalidixic acid, ciprofloxacin, gentamicin, and chloramphenicol. In addition, the R-pyocin susceptibility pattern was determined and the strains were O-serotyped and tested for their agglutination in acriflavine. The isolates included 53 strains resistant to both gentamicin and carbenicillin, 13 to carbenicillin only, 20 to gentamicin only, and 30 sensitive to gentamicin and carbenicillin. The multiresistant isolates displayed relatively higher resistance to all other antibiotics except aztreonam and cefotaxime. Remarkably 30 out of 53 multiresistant isolates reacted with one pyocin only, namely pyocin R2. This R-pyocin response was not encountered in any other strains of the other antibiotic resistance phenotypes. These isolates belonged to the 0-12 serogroup. The 0-12 serogroup was represented only in a minority of strains giving other R-pyocin reactions. It is interesting that strains reacting with pyocin R5 only were mostly susceptible to antibiotics. The results clearly indicate lipopolysaccharide-core mutations in multiresistant clinical isolates of P. aeruginosa. Despite the fact that the R-pyocin resistance pattern can not define the precise possible defect, the multiple and high level resistance associated with R2-pyocin reaction seems to be an interesting trait.     

Modification of Pseudomonas aeruginosa virulence factors by sub-inhibitory concentrations of antibiotics.

This study's objectives were to evaluate the effects of subminimum inhibitory concentrations (MICs) of tobramycin, gentamicin, netilmicin, streptomycin, ciprofloxacin, cefotaxime and piperacillin on proteinase production, alginate and siderophore synthesis by two strains of Pseudomonas aeruginosa. One of these strains, of recent clinical isolation, was mucoid. In fact it is well known that mucoid strains are more resistant than non-mucoid; there is, moreover, evidence that in cystic fibrotic lungs the non-mucoid P. aeruginosa are invariably replaced by mucoid variants. Our results show that subinhibitory concentrations of beta-lactams and quinolones significantly reduced the amount of alginate. Protease production was affected by all antibiotics tested.     

Modification of Pseudomonas aeruginosa Virulence Factors by Sub-Inhibitory Concentrations of Antibiotics

This study’s objectives were to evaluate the effects of subminimum inhibitory concentrations (MIC_s) of tobramycin, gentamicin, netilmicin, streptomycin, ciprofloxacin, cefotaxime and piperacillin on proteinase production, alginate and siderophore synthesis by two strains of Pseudomonas aeruginosa. One of these strains, of recent clinical isolation, was mucoid. In fact it is well known that mucoid strains are more resistant than non-mucoid; there is, moreover, evidence that in cystic fibrotic lungs the non-mucoid P. aeruginosa are invariably replaced by mucoid variants. Our results show that subinhibitory concentrations of β-lactams and quinolones significantly reduced the amount of alginate. Protease production was affected by all antibiotics tested.     

Relationship of Antibiotic Resistance Phenotype to the R-Pyocin Susceptibility Pattern in Clinical Isolates of Pseudomonas aeruginosa

Summary

One hundred sixteen clinical isolates of Pseudotnonas aeruginosa were collected from 7 hospitals in Athens. All strains were studied for their susceptibility to cefotaxime, ceftazidime, carbenicillin, aztreonam, imipenem, nalidixic acid, ciprofloxacin, gentamicin, and chloramphenicol. In addition, the R-pyocin susceptibility pattern was determined and the strains were O-serotyped and tested for their agglutination in acriflavine. The isolates included 53 strains resistant to both gentamicin and carbenicillin, 13 to carbenicillin only, 20 to gentamicin only, and 30 sensitive to gentamicin and carbenicillin. The multiresistant isolates displayed relatively higher resistance to all other antibiotics except aztreonam and cefotaxime. Remarkably 30 out of 53 multiresistant isolates reacted with one pyocin only, namely pyocin R2. This R-pyocin response was not encountered in any other strains of the other antibiotic resistance phenotypes. These isolates belonged to the 0–12 serogroup. The 0–12 serogroup was represented only in a minority of strains giving other R-pyocin reactions. It is interesting that strains reacting with pyocin R5 only were mostly susceptible to antibiotics. The results clearly indicate lipopolysaccharide-core mutations in multiresistant clinical isolates of P. aeruginosa. Despite the fact that the R-pyocin resistance pattern can not define the precise possible defect, the multiple and high level resistance associated with R2-pyocin reaction seems to be an interesting trait.     

In vitro effect of subminimal inhibitory concentrations of antibiotics on the biofilm formation ability of Acinetobacter baumannii clinical isolates

The ability of A cinetobacter baumannii strains to form biofilm is one of the most important virulence factor which enables bacterial survival in a harsh environment and decreases antibiotic concentration as well. Subminimal inhibitory concentrations (subMICs) of antibiotics may change bacterial ultrastructure or have an influence on some different molecular mechanisms resulting in morphological or physiological changes in bacteria itself. The aim of this study was to determine effects of 1/2, 1/4, 1/8 and 1/16 minimal inhibitory concentrationsof imipenem, ampicillin-sulbactam, azithromycin, rifampicin and colistin on biofilm formation ability of 22 biofilm non-producing and 46 biofilm producing A. baumannii strains (30 weak producing strains and 16 moderate producing strains). Results of this study indicate that 1/2–1/16 MICs of imipenem, azithromycin, and rifampicin can reduce bacterial biofilm formation ability in moderate producing strains (p < 0.05), whereas 1/16 MIC of imipenem and 1/4–1/8 MICs of rifampicin reduce the biofilm formation in weak producing strains (p < 0.05). Statisticaly significant effect was detected among biofilm non-producing strains after their exposure to 1/16 MIC of azithromycin (p = 0.039). SubMICs of ampicillin-sulbactam and colistin did not have any significant effect on biofilm formation among tested A. baumannii strains.     

Resistance to antibiotics and inorganic ions in virulent bacterial strains from a hospital

Virulent strains of Staphylococcus aureus, S. epidermidis, Escherichia coli and Pseudomonas aeruginosa were studied for their resistance to antibiotics and inorganic ions, the correlation with their clinical use and the usefulness as an epidemioliogical tool. Multiresistance was common, the antibiotypes were similar to those previously reported, but characteristic resistotypes endemic of our county were found. A correlation between resistance and metal ion consumption was not detected. Staphylococci strains were susceptible to vancomycin, cephalothin and mercury chloride; S. epidermidis showed higher rates of resistance to antibiotics and lower to cadmium chloride and potassium iodine than S. aureus. E. coli strains were susceptible to new beta-lactamans; resistance to cephalothin, gentamicin, tobramycin and amikacin was less than 10%. P, aeruginosa was the species with the most multiresistance and antibiotype diversity, only ceftazidime, amikacin and imipenem had a resistance rate less than 11%. In both E. coli and P. aeruginosa resistance to all tested metals (except silver nitrate) was found although with different percentages.     

Evaluation of different compounds as quorum sensing inhibitors in Pseudomonas aeruginosa

Quorum sensing (QS) controls systems affecting the pathogenicity of many microorganisms; its interruption has an anti-pathogenic effect and can be used in the treatment of bacterial infections. In this study we evaluated QS regulation by Pseudomonas aeruginosa strains and QS inhibition (QSI) by different compounds. The inhibitory activity of 3 macrolide and 3 lincosamide drugs, resveratrol, garlic extract and N-acetylcysteine was tested on 4 P. aeruginosa strains isolated from cystic fibrosis (CF) patients using Chromobacterium violaceum ATCC 12472 as biomonitor. One P. aeruginosa strain, lincomycin and N-acetylcysteine did not show QSI, contrary to other compounds and P. aeruginosa strains. These results indicate that QSI evaluation should be taken into account in the design of new therapeutic strategies to treat P. aeruginosa infections, especially in patients infected by antibiotic-resistant bacteria.     

Anticandidal activity of Pseudomonas aeruginosa strains isolated from clinical specimens

Pseudomonads represent the major group of non-differentiating microorganisms that produce antibiotics. The antibiotic substances produced by this group of organisms are pyocyanin, pyrolnitrin and pseudomonic acid. This study was designed to investigate the in vivo and in vitro anticandidal activity of Pseudomonas aeruginosa strains against Candida species. Forty-four P. aeruginosa strains isolated from various specimens of intensive care patients were included in the study. All P. aeruginosa strains have pyocyanin pigment. Candida albicans ATCC 10231, Candida parapsilosis ATCC 22019, Candida krusei ATCC 6258 and a clinical isolate of Candida tropicalis were used to measure the anticandidal activity of Pseudomonas strains by Kerr's method. The total inhibition rates obtained by using blood agar of C. albicans, C. parapsilosis, C. krusei and C. tropicalis were 41%, 34%, 34% and 25% respectively. When Sabouraud dextrose agar (SDA) was used, the rates were detected as 45%, 39%, 48% and 25% respectively. In the mouse model of concomitant subcutaneous infection with Candida species and P. aeruginosa no yeast were recovered from skin cultures despite 100% detection of P. aeruginosa. Pseudomonas aeruginosa strains isolated from intensive care patients showed anticandidal activity against the Candida species in the present study and this point may be important in the following and treatment of patients.     

Enhanced in-vitro activity of liposome-trapped penicillin-G against Pseudomonas aeruginosa

The growth inhibition of four Pseudomonas aeruginosa strains by liposome-trapped penicillin-G was investigated. There were indications of an association of the efficacy of liposomal penicillin-G with the nature of the 0-antigenic polymeric side chain. Namely, P28-800 and PCF-95 strains, characterized by a rough polysaccharide chain, were the most susceptible, whereas strain P28-0, possessing an intact lipopolysaccharide, resisted the activity of the entrapped drug. Among the rough strains, P642, a beta-lactamase producer, was not affected by the encapsulated drug. The composition of liposomes seems to have a significant impact in arresting the growth of the P. aeruginosa strain.     

Widespread detection of VEB-1-type extended-spectrum beta-lactamases among nosocomial ceftazidime-resistant Pseudomonas aeruginosa isolates in Sofia, Bulgaria

A total of 132 ceftazidime-resistant clinical isolates of Pseudomonas aeruginosa were collected during 2001-2005 from 5 university hospitals in Sofia, Bulgaria to assess the current levels of antimicrobial susceptibility and to evaluate resistance mechanisms to beta-lactams. Antimicrobial susceptibilities were detected by a disk diffusion method and E-test. Polymerase chain reaction amplification and sequencing of bla(VEB-1 )and bla(PER-1 )were performed. The antibiotic resistance rates were: to piperacillin 90.2%, piperacillin/tazobactam 52.3%, ceftazidime 94.7%, cefepime 88.6%, cefpirome 98.5%, aztreonam 85.6%, imipenem 66.6%, meropenem 63.6%, amikacin 81.1%, gentamicin 84.8%, tobramycin 89.4%, netilmicin 57.6%, ciprofloxacin 83.4%. Structural genes for VEB-1 extended-spectrum beta -lactamases (ESBLs) were found in 75 (56.8%) of the isolates. PER-1 ESBLs were not detected. The VEB-1-producing strains were more resistant than VEB-1 non-producers to amikacin, gentamicin, tobramycin and ciprofloxacin ( P<0.001). VEB-1 appears to have a significant presence among ceftazidime-resistant P. aeruginosa isolates from Sofia.     

Phage F-116 Transduction of Antibiotic Resistance from a Clinical Isolate of Pseudomonas aeruginosa

Summary

The lysate of phage F-116, propagated in a multiple drug resistant clinical isolate of Pseudomonas aeruginosa No. 131 was used to transduce determinants of antibiotic resistance to susceptible auxotrophic laboratory strains of the same species. The phage preparation, designated F-116/131 was found to transduce four determinants of resistance, i.e. to imipenem, cefotaxime, kanamycin and carbenicillin, but not to streptomycin, gentamicin, ceftazidime nor ciprofloxacin/ofloxacin. No conjugal transfer of any resistance determinants could be demonstrated in mating experiments using strain No. 131 and two rifampicin-resistant strains of P. aeruginosa which were highly susceptible to all antibiotics studied. These results might suggest that transduction could be an additional way to conjugational transfer of antibiotic resistance among P. aeruginosa.     

Transferable Resistance to Imipenem in Hospital Isolates of Pseudomonas aeruginosa

Summary

We monitored systematically, for more than five years, the eventual transferability of resistance to imipenem in strains of Pseudomonas aeruginosa isolated from patients in Frankfurt University Clinics. Quite recently, four strains have been found which transfer resistance to imipenem to recipient strains of P. aeruginosa. Although in three strains imipenem was the only antibiotic where resistance was transferred directly, the indirect selection analysis showed that, in each instance, determinants of resistance to carbenicillin and kanamycin were co-transferred. The situation in the fourth strain was more complicated. It was resistant to at least ten anti-pseudomonad antibiotics, and transferred directly not only determinants of resistance to imipenem, but also to carbenicillin and kanamycin, as did the other strains, plus determinants of resistance to ceftazidime and cefotaxime. The origin and mode of spread of resistance determinants in studied strains is briefly discussed.     

A retrospective study of Pseudomonas aeruginosa isolation from clinical sources

The incidence of Pseudomonas aeruginosa recovery from blood, urine and other clinical specimens as well as its resistance to various antimicrobials during a 5-year period (1982 to 1986) are reported. The susceptibility patterns to newer antipseudomonad agents of 100 randomly selected P. aeruginosa clinical strains isolated in 1987 is also presented. Our results show an overall P. aeruginosa isolation incidence of 10.6%, with its resistance to routinely used antibiotics being high but relatively low to the newer agents which have not been used extensively in our hospital.     

Antiseptic effect of a novel alcohol-free mouthwash: a convenient prophylactic alternative for high-risk patients

We developed an efficacious and non-irritant mouthwash that is alcohol-free and that has a low concentration of chlorhexidine, in order to be used for preventing oral cavity infections in immunocompromised and cancer patients. The novel mouthwash solution was tested for its antimicrobial efficacy against both free floating (planktonic) and the biofilm forms of Candida albicans. The solution was also tested against Klebsiella pneumoniae, Pseudomonas aeruginosa, and methicillin-resistant Staphylococcus aureus (MRSA), using a modification of a previously published method. The activity of the novel mouthwash was also compared with that of three commercially available mouthwashes. The experimental mouthwash showed efficacy against C. albicans, both in free-floating form and in biofilm. With higher concentration of chlorhexidine, the solution was also efficacious in inhibiting the growth of K. pneumoniae, P. aeruginosa, and MRSA. The antiseptic activity of the alcohol-free mouthwash against other bacterial organisms and C. albicans was comparable to other commercially available alcohol-based mouthwash solutions. A novel alcohol-free mouthwash solution, that has low concentration of chlorhexidine, showed antiseptic effect against planktonic and biofilm forms of C. albicans and against K. pneumoniae, P. aeruginosa, and MRSA.     

In vitro evaluation of the antimicrobial activity of meropenem in combination with polymyxin B and gatifloxacin against Pseudomonas aeruginosa and Acinetobacter baumannii

The antimicrobial activity of meropenem combined with either polymyxin B or gatifloxacin was evaluated by the checkerboard method against Pseudomonas aeruginosa (10 strains) and Acinetobacter baumannii (10 strains). In addition, the triple combination of polymyxin B, gatifloxacin, and meropenem was also studied as well as the polymyxin B and gatifloxacin combination. A partial synergism interaction between meropenem and polymyxin B was observed for 80% of the A. baumannii strains. In contrast, this combination showed an indifferent effect for 80% of the P. aeruginosa strains tested. The combination of meropenem and gatifloxacin showed synergism only for two strains of A. baumannii, and partial synergism and additive effect for seven strains and indifference for four strains of both species. For the strains of P. aeruginosa, the double combination of polymyxin B and gatifloxacin and the triple combination of meropenem, polymyxin B and gatifloxacin were indifferent for the majority of the strains tested, that is, 90 and 80% respectively.     

Antibiotic susceptibility tests directly on urine samples using Uro-Quick, a rapid automated system

During the period June 2003-March 2004, 12,579 urine samples were examined employing the Uro-Quick system. Positive samples (1,948) were subsequently Gram-stained and processed by standard procedures for microorganism identification and antibiotic susceptibility determination by the disk diffusion method. Results of this latter test were compared with those obtained employing the new rapid Uro-Quick method. Antibiotics were introduced in vials containing 2 ml of Mueller-Hinton broth, then 0.5 ml of urine or a bacterial suspension in broth were added; a vial without drug was used as control. After 3 and 5 hours of incubation (for Gram-negative and Gram-positive strains respectively) the instrument showed the results. No growth and a growth curve like the control indicated susceptible and resistant strains respectively. Overall 1,590 Gram-negative strains were tested against ciprofloxacin, nitrofurantoin, amoxicillin-clavulanate, ceftazidime, fosfomycin, imipenem, amikacin, trimethoprim-sulfamethoxazole, and piperacillin-tazobactam, while 358 Gram-positive bacteria were assessed against ciprofloxacin, nitrofurantoin, amoxicillin-clavulanate, ampicillin, fosfomycin, gentamicin, oxacillin and trimethoprim-sulfamethoxazole. Against the major urinary tract pathogens (Escherichia. coli, enterococci, Klebsiella spp. and Proteus spp.) agreement between the Uro-Quick system and the disk diffusion test generally was >90% for all antibiotics tested. On the basis of these results the system appears useful not only for bacteriuria screening, but also to rapidly test the antibiotic susceptibility of common uropathogens.     

Antibiotic resistance among nosocomial isolates in a Croatian intensive care unit--results of a twelve-year focal surveillance of nosocomial infections

Continuous 12-year (1990--2001) focal surveillance of the antibiotic resistance among the most common nosocomial pathogens (Klebsiella pneumoniae, Pseudomonas aeruginosa, Acinetobacter sp., and Staphylococcus aureus) in 1325 Intensive Care Unit patients was performed. The surveillance period was divided in three 4-year time intervals (1990--1993, 1994--1997 and 1998--2001) and the prevalence of resistance was compared between intervals. Specimens included blood, urine and respiratory tract specimens. The incidence and trends of resistance to six antibiotics showed inconsistent results. Aminoglycoside resistance decreased among K. pneumoniae_isolates (gentamicin 83%, 72.7% and 49.6%; amikacin 50.9%, 51.5% and 18.2%) and Acinetobacter sp. strains (amikacin 77%, 63.4% and 58.2%) but increased in P. aeruginosa (amikacin 27.5%, 63.3% and 44.1%). Overall, resistance to ceftazidime, ciprofloxacin, and imipenem increased but imipenem resistance is still low, particularly among Acinetobacter sp. isolates (0, 2.1% and 1.5%). However, imipenem resistance increased among P. aeruginosa (10.2%, 31.6%, 22.1%). The prevalence of methicillin resistance was high but did not change during the surveillance period (82.3%, 78.3% and 82.2%). The present study suggests a complex picture of the development of antibiotic resistance in a single ICU. Significant changes occur over time but they are unpredictable and do not show identical tendencies for different species and antibiotics.