Retrospective analysis of antibiotic susceptibility patterns of respiratory isolates of Pseudomonas aeruginosa in a Turkish University Hospital

Background Lower respiratory tract infections due to Pseudomonas aeruginosa have a high mortality rate. Antibacterial activity of various antibiotics against P. aeruginosa isolated from each hospital depends on the variety or amount of antibiotics used in each hospital. Method A total of 249 respiratory isolates of Pseudomonas aeruginosa in Sivas (Turkey) were included between January-1999 and January-2002. Isolates were tested against 14 different antibiotics by a disc diffusion method or standardized microdilution technique. Results Organisms were cultured from the following specimens: sputum (31.3%), transtracheal/endotracheal aspirates (37.8%), and bronchial lavage (30.9%). Isolates in bronchial lavage were highly susceptible to cefoperazone and aminoglycosides. Resistance to ampicillin/sulbactam was 98.8%, ticarcillin 40.1%, ticarcillin/clavulanic acid 11.2%, piperacillin 21.8%, aztreonam 66.6%, cefotaxim 75.4%, ceftriaxone 84.2%, cefoperazone 39.0%, ceftazidime 50.8%, gentamicin 57.5%, tobramycin 58.4%, amikacin 25.4%, ciprofloxacin 16.1%, and imipenem/cilastatin 21.6%. The term multidrug-resistant P. aeruginosa covered resistance to imipenem, ciprofloxacin, ceftazidime, gentamicin, and piperacillin. 1.2% of isolates were multidrug-resistant. Conclusions These findings suggest that amikacin resistance increases progressively in Turkey. Piperacillin and ticarcillin/clavulanate were the most active agents against both imipenem- and ciprofloxacin-resistant isolates in our region.


Background
Pseudomonas aeruginosa is primarily a nosocomial pathogen and the most common gram-negative bacillus causing hospital-acquired pneumonia. Despite therapy, the mortality from hospital-acquired pseudomonal pneumonia is approximately 70 percent [1]. Unfortunately, there are no specific measures to prevent nosocomial pseudomonal infections. Despite the availability of a variety of effective antimicrobial agents, treatment of pseudomonal pneumonia is often challenging. The aim of this study was to reveal the antibiotic susceptibility patterns of Pseudomonas aeruginosa isolated airway samples in Sivas, Central Anatolia.

Material and methods
The antibiotic susceptibility patterns of 249 respiratory isolates of Pseudomonas aeruginosa were retrospectively analyzed. Data between 01.01.1999 and 01.01.2002 was obtained in microbiology laboratory of University of Cumhuriyet and processed to eliminate duplicate registrations. The isolates were identified conventional or automatized (Api) techniques. Isolates collected from the same specimen source of the same patient within 7 days were excluded. Organisms were cultured from the following specimens: sputum (31.3%), transtracheal/endotracheal aspirates (37.8%), and bronchial lavage (30.9%). All consecutive isolates were accepted, as we did not attempt to distinguish the actual pathogens from colonizing strains.
Susceptibility to 14 antimicrobial agents was confirmed at the central laboratory by disk diffusion according to the National Committee for Clinical Laboratory Standards guidelines [2]. Inoculated plates were incubated aerobically at 35°C and estimated after 18 h. For quality control of the disk diffusion tests E. coli ATCC 25922 and S. aureus ATCC 25923 strains were used. Disk diffusion method was used until November 1999 in our microbiology laboratory, and later antibiotic resistance patterns were analyzed by a co-ordinating laboratory that determined minimal inhibitory concentrations of these 14 antimicrobial agents using a standardized microdilution technique (Sceptor system, Becton Dickinson Microbiology System).
The results of the susceptibility testing were classified into two categories. The category "susceptible" was defined as identification of a strain as susceptible by the disk diffusion method or microdilution technique. All resistant and intermediate isolates of the species were classified under the definition "resistant".
The significance of differences in resistance was evaluated using EpiInfo software (version 5.0). Susceptibility and resistance were calculated as percentages with 95% confidence intervals. The X 2 test with Yates correction or Fisher's exact two-tailed test was used to assess the association of selected variables with the prevalence of resistant isolates. The analysis was performed on the cross-tabulated values of the presence of the resistant/susceptible isolates, according to the categories of the selected variable. A p-value of <0.05 was considered statistically significant.
There was not significant difference in the susceptibility to ureidopenicillins in imipenem-, gentamicin-, and ciprofloxacin-resistant strains compared to the entire group. Piperacillin-resistant strains were more resistant to aztreonam, ceftazidime, ticarcillin, and ticarcillin/clavulanate than the entire group. Piperacillin-resistant strains were more susceptible to cefotaxim and tobramycin but, more resistant to ticarcillin/clavulanate than gentamicin-resistant strains (p < 0.05).

Discussion
Resistance of gram-negative aerobic bacteria to aminoglycoside antibiotics differs by region and country. Resistance to aminoglycosides was higher in Southern Europe than in Central and Northern Europe. Reports of the susceptibility of P. aeruginosa to gentamicin and tobramycin have ranged from as low as 49.8% and 77.7%, in Greece, to as high as 96.6% and 99.2%, respectively, in the United Kingdom [3]. It was reported that 54% of gram-negative bacilli in Turkey are resistant to gentamicin, 35% to tobramycin, and only 0.9% to amikacin in 1988 [4]. Consistent to this finding, resistance to amikacin (25.4%) of P. aeruginosa was still lower than to gentamicin (57.5%) or tobramycin (58.4%). However, this data suggests that resistance to amikacin increases progressively in Turkey.
Isolates in tracheal aspirates were highly resistant whereas isolates obtained from bronchial lavage fluids were relatively quite susceptible. In our hospital, tracheal aspiration is generally a procedure performed in intensive care units or the patients with critical illness. Consequently, our isolates in tracheal aspirates may reflect nosocomial strains. On the other hand, bronchoscopy, a lesser invasive procedure, was generally performed to newly hospitalised patients. The variations of antibiotic resistance for different locations may be due to the isolation procedures.
Isolates that were resistant to one class of antibiotics were also resistant to at least one other class of antibiotics except resistance to piperacillin. This finding suggests that piperacillin resistance occur independently and specifically in our isolates compared to other anti-pseudomonal agents. One hundred isolates were resistant to ticarcilin but 81 of them were susceptible to ticarcillin-clavulanate, which suggests that the production of β-lactamase may be the major mechanism of resistance in these organisms.
Gaynes and Culver demonstrated that resistance was more common among P. aeruginosa isolated from the respiratory tract, patients in intensive care units, and in teaching hospitals [10]. In spite of this knowledge, it was found only three multidrug-resistant isolates in 3-year survey. We think that our rate of multidrug-resistance (1.2%) was reasonable.

Conclusions
Because of high resistance rates to extended-spectrum cephalosporins and the progression of amikacin resistance, a national strategy on the limited and prudent use of antipseudomonal agents is urgently needed in Turkey.