A contemporary cohort study of consecutive patients with P. aeruginosa nosocomial infections was performed at two tertiary-care teaching hospitals in Porto Alegre, southern Brazil. The study period was from September 2004 to June 2005 at São Lucas Hospital, a 600-bed hospital, and from January to June 2005 at Hospital de Clínicas de Porto Alegre, a 1,200-bed hospital 7.

In the current study, we analyzed patients ≥ 18 years, who did not have cystic fibrosis, who had been diagnosed with HAP defined as follows. First, the presence of positive cultures for P. aeruginosa either recovered from respiratory secretions (>10⁶ cfu/ml from endotracheal aspirates or >10⁴ cfu/ml from bronchoalveolar lavage) after 48 hours of hospital admission, or within 48 hours if the patient had been hospitalized in the past 60 days, or recovered from blood without the presence of any other pathogen in respiratory secretions. Second, the presence of a radiographic infiltrate that was new or progressive, along with the presence of two or more of the following criteria: fever (temperature >38°C) or hypothermia (temperature <36°C), purulent sputum, leukocytosis (>10,000 cells/mm³) or leukopenia (<4,000 cells/mm³), and a decline in oxygenation. Sputum was considered purulent if >25 neutrophiles and <10 epithelial cells per high power field were present. VAP was defined as HAP that developed after 48 hours of mechanical ventilation.

Patients were excluded if they did not fulfill these criteria for HAP. Patients were followed from the first isolation of P. aeruginosa to discharge from hospital or to death. Antimicrobial agents used were at the discretion of the patient's physicians, not the investigators. The ethics review boards of both hospitals approved the study.

Data were collected from medical charts and/or hospital computer system databases, both during and after the patients' hospitalization. The researchers were blinded for the MBL status of P. aeruginosa isolates during data collection.

Conventional microbiology methods were used for P. aeruginosa identification, and susceptibility tests were performed by disk-diffusion methods according to Clinical and Laboratory Standards Institute, (formerly National Committee for Clinical Laboratory Standards), guidelines 8. Susceptibility was tested for amikacin, aztreonam, cefepime, ceftazidime, ciprofloxacin, imipenem, meropenem, piperacillin-tazobactam, and polymyxin B. Susceptibility of the latter was determined using the interpretative criteria (≥ 14 mm) proposed elsewhere 9. All isolates resistant to ceftazidime were screened for MBL production with ceftazidime in the presence of 3 μl 2-mercaptopropionic acid as previously described 10.

The main outcome was 30-day mortality. Other secondary outcomes were the length of need for vasoactive drugs and the length of mechanical ventilation (both were assessed in survivors).

The variable in the study was MBL production. Other independent variables analyzed included the following: age; sex; Charlson comorbidity score 11 (assessed at the moment of HAP diagnosis); baseline diseases; iatrogenic immunosuppression, such as chemotherapy-induced neutropenia (neutrophile count ≤ 1,000 mm³), and/or receipt of corticoid drugs (prednisone ≥ 10 mg daily or equivalent doses) or other immunosuppressive agents for >14 days; the presence of other concomitant infections (infections by other organisms at a site other than the lung, excluding coagulase-negative staphylococci in a single blood culture); a previous surgical procedure during the hospital stay; the length of hospital stay (before the diagnosis of HAP); presentation of HAP with severe sepsis or septic shock 12; infection by P. aeruginosa at more than one site (not including patients with HAP and bacteremia); polymicrobial infection (isolation of another organism from the respiratory secretions at the moment of P. aeruginosa HAP diagnosis); associated bacteremia (isolation of P. aeruginosa from one or more blood samples); VAP; receiving appropriate empirical therapy (defined as the administration of an antimicrobial agent to which the isolate was susceptible in vitro in ≤ 24 hours of sample collection); receiving appropriate definitive therapy (defined as the use for at least 48 hours of an antimicrobial agent to which the isolate was susceptible in vitro); time to receiving appropriate definitive therapy (only for those who have not received appropriate empirical therapy; time in days from the sample collection to the first dose of appropriate therapy); and combination antibiotic treatment (treatment with more that one agent with in vitro susceptibility).

Aminoglycosides in monotherapy were not considered appropriate treatment therapy despite in vitro susceptibility 3.

All statistical analyses were carried out using SPSS for Windows, version 13.0. The relative risk (RR) and the 95% confidence interval (CI) were calculated for 30-day mortality of patients with MBL-PA HAP and of patients with non-MBL-PA HAP. P values were calculated using the chi-squared test or Fischer exact test for categorical variables, and using Student's t test or the Wilcoxon rank-sum test for continuous variables.

A logistic regression model was constructed to identify independent factors associated with 30-day mortality using a forward stepwise approach. Variables for which the P value was < 0.20 in univariate analysis were included in the model. P = 0.05 was set as the limit for acceptance or removal of the terms in the model. MBL production remained in the model independent of the P value. All tests were two-tailed and P ≤ 0.05 was considered significant.

A total of 473 patients presented the isolation of P. aeruginosa after >48 hours of hospital admission. Of these, 171 presented the isolation of P. aeruginosa in respiratory secretions. Twenty-one patients were excluded because they did not fulfill the criteria for HAP. A total of 150 patients were analyzed. Forty-two (28.0%) patients presented MBL-PA HAP.

The 30-day mortality was 37.3% (56 of 150) and represented 76.7% of the 73 deaths. Among patients with MBL-PA HAP the 30-day mortality was 57.1% (24 of 42), compared with 29.6% (32 of 108) for non-MBL-PA patients (RR, 1.93; 95% CI, 1.30–2.85; P < 0.01). The overall mortality rate was 18.7 per 1,000 patient-days: 26.5 per 1,000 patient-days among MBL-PA-infected patients and 15.8 per 1,000 patient-days among non-MBL-PA-infected patients (P = 0.02). The median length of follow-up was 19 days (interquartile range, 9–31 days): 16 days (interquartile range, 7–28 days) for those patients with MBL-PA HAP and 19.5 days (interquartile range, 10–31.5 days) for those patients with non-MBL-PA HAP (P = 0.19). The median length of follow-up of those patients who did not die within 30 days did not differ between patients with MBL-PA HAP and patients with non-MBL-PA HAP (29.5 days (interquartile range, 22–71 days) versus 24.5 days (interquartile range, 14–39.5 days), respectively; P = 0.22).

Fifty-five patients (36.7%) had VAP. Patients with VAP had a 30-day mortality of 60.0% (33 of 55): 77.3% (17 of 22) for patients with MBL-PA VAP compared with 48.5% (16 of 33) for those patients with non-MBL-PA VAP (RR, 1.59; 95% CI, 1.05–2.42; P = 0.03).

The characteristics of patients according to 30-day mortality are presented in Table 1. Factors associated with mortality within 30 days in the univariate analysis were severe sepsis or septic shock, VAP, and bacteremia. Comorbidity scores were higher in patients who died within 30 days, but there was no statistically significant difference. Both empirically appropriate therapy and receiving appropriate therapy at any moment were significant protective factors for 30-day mortality; however, a greater effect was observed for therapy at any time.

Considering only patients who received appropriate therapy (n = 109), there was no statistically significant difference in mortality rates according to the time to administration of appropriate therapy. The 30-day mortality rates were 21.7% (≤ 24 hours), 32.0% (>24 hours but ≤ 72 hours), and 34.2% (>72 hours) (P = 0.41).

The results of multivariate analysis are presented in Table 2. The Charlson score, severe sepsis or septic shock, VAP, and appropriate treatment at any moment were significantly associated with 30-day mortality. Bacteremia and antimicrobial combination were not statistically significant and were excluded from the model. MBL production was not significantly associated with the outcome in the final model, but was statistically significant in the multivariate model (RR, 2.84; 95% CI, 1.24–6.52, P = 0.01) before the inclusion of appropriate antimicrobial therapy in the model. Specific comorbidities such as cirrhosis and AIDS were not included in the model because they were significantly associated with higher Charlson scores (data not shown).

Among the 77 survivors, patients with MBL-PA HAP presented significantly longer length of need for vasoactive drug therapy than non-MBL-PA patients (mean, 17.5 ± 3.5 days versus 3.6 ± 3.3 days; P < 0.001). The length of mechanical ventilation was also longer for patients with MBL-PA HAP than for non-MBL-PA patients, although without statistical significance (mean, 13.0 ± 9.0 days versus 7.8 ± 4.9 days; P = 0.12).

A total of 38 distinct antibiotic resistance profiles were observed in P. aeruginosa isolates, but only six distinct patterns were observed among MBL-PA isolates. These latter profiles are presented in Table 3. Among non-MBL-PA isolates, 36 resistance profiles were found. The commonest profile was susceptibility to all tested drugs (27 patients, 25.0%), followed by susceptibility to all drugs except aztreonam (14 patients, 13.0%), and susceptibility to ceftazidime and piperacillin-tazobactam and resistance to the other drugs (10 patients, 9.3%). Other profiles each accounted for 6.5% or less of the total.

Among the 42 patients with MBL-PA HAP, 41 received antimicrobial therapy and one did not receive any antibiotic. This latter patient had a missed diagnosis of HAP and died after four days of the onset of infection. Twenty-one (51.2%) of the 41 treated patients received appropriate therapy: 10 patients (47.6%) received such therapy in ≤ 72 hours, and three patients (14.3%) received appropriate antibiotic in ≤ 24 hours. Patients who received any therapy in ≤ 72 hours (n = 10) tended to present a lower 30-day mortality than those who received therapy after 72 hours (n = 31): 30.0% and 64.5%, respectively (RR, 0.45; 95% CI, 0.17–1.24; P = 0.06). Antimicrobial therapies of patients with MBL-PA HAP are presented in Table 4.

Considering only those 21 patients who received appropriate therapy, there was no statistically significant difference in mortality between receiving therapy in ≤ 72 hours (n = 9) and in >72 hours (n = 12) for 30-day mortality (33.3% versus 50.0%; RR, 0.67; 95% CI, 0.23–1.97; P = 0.38). Among these patients, no specific antibiotic agent was associated with lower mortality (P = 0.54).