Comparison of MALDI-TOF MS and AFLP for strain typing of ESBL-producing Escherichia coli.

Typing of bacterial isolates using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) potentially provides an efficient on-site method to monitor the spread of antibiotic-resistant bacteria and rapidly detect outbreaks. We compared MALDI-MS typing results to those of amplified fragment length polymorphism (AFLP) in a collection of 52 ESBL-producing Escherichia coli, isolated in a Dutch nursing home with an on-going outbreak of ST131 E. coli. Specific MALDI types were defined based on spectral data from four replicate colony samples of isolates grown on Columbia agar using multivariate statistical procedures. Type-specific superspectra were computed for four E .coli MALDI-types and tested for the potential of rapid and automated typing. The effect of different incubation conditions on typing performance was tested by analysing five isolates incubated for 24 h and 48 h on five different media. Types defined based on MALDI spectra were largely in agreement with the AFLP results, although some MALDI types comprised of more than one AFLP type. In particular, isolates belonging to ST131 showed distinct mass patterns. The proportion of isolates correctly assigned was substantially lower for isolates incubated on Sabouraud-dextrose and Drigalski agars for 24 h, and for those incubated for 48 h (all media). Our results show that the identification of type-specific peaks potentially allows direct typing of isolates belonging to specific clonal lineages. Both incubation time and media affected type assignment, suggesting that there is a need for a careful standardization of incubation time and culturing conditions when developing MALDI-typing schemes for E. coli.

Community-acquired methicillin-resistant Staphylococcus aureus: current perspectives.

Methicillin-resistant Staphylococcus aureus (MRSA) has become a worldwide problem, although its prevalence varies considerably among countries. The epidemiology of MRSA is now changing; infections are no longer confined to the hospital setting, but also appear in healthy community-dwelling individuals without established risk factors for the acquisition of MRSA. Reported prevalence rates of community-acquired MRSA (CA-MRSA) vary widely among studies-largely because of the different definitions employed and different settings in which the studies have been performed. At present, molecular epidemiological definitions, based on staphylococcal cassette chromosome mec (SCCmec) typing and phylogenetic analyses of the MRSA isolates, are considered the most reliable means by which to distinguish between hospital-acquired MRSA (HA-MRSA) and CA-MRSA. CA-MRSA has been isolated predominantly from skin and soft tissue infections, such as abscesses, cellulitis, folliculitis and impetigo. Although CA-MRSA infections are usually mild, they may also be severe, and can result in hospitalisation and even death. CA-MRSA strains differ from the major pandemic clones of MRSA that account for the majority of epidemic HA-MRSA strains. Differences are found in SCCmec types, bacterial growth rate, and the distribution of antibiotic resistance genes and toxin genes. Mathematical models have shown that CA-MRSA has a high potential to become endemic in the community, and this will impact significantly on the control of MRSA in the hospital setting. Well-designed, community-based studies with adequate risk factor analysis are required to further elucidate the epidemiology of CA-MRSA and to improve strategies to control MRSA in both the community and hospital settings.

Vancomycin versus placebo for treating persistent fever in patients with neutropenic cancer receiving piperacillin-tazobactam monotherapy.

This prospective, double-blind trial assessed whether the addition of a glycopeptide would be able to reduce the time to defervescence in neutropenic patients with cancer who had persistent fever 48-60 h after the initiation of empirical piperacillin-tazobactam monotherapy. Of 763 eligible patients, 165 with persistent fever were randomized to receive piperacillin-tazobactam therapy plus either vancomycin therapy or placebo. Defervescence was observed in 82 (95%) of 86 patients in the vancomycin group and in 73 (92%) of 79 patients in the placebo group (P=.52). The distributions of the time to defervescence were not statistically significant between the 2 groups (estimated hazard ratio, 1.03; 95% confidence interval, 0.75-1.43; P=.75). The number of additional episodes of gram-positive bacteremia and the percentage of patients for whom amphotericin B was empirically added to their therapy regimen were also similar in both groups. This study failed to demonstrate that the empirical addition of vancomycin therapy to the treatment regimen is of benefit to persistently febrile neutropenic patients with cancer.

Reduction of surgical-site infections in cardiothoracic surgery by elimination of nasal carriage of Staphylococcus aureus.

OBJECTIVE: To test the hypothesis that perioperative elimination of nasal carriage of Staphylococcus aureus using mupirocin nasal ointment reduces the surgical-site infection (SSI) rate in cardiothoracic surgery. DESIGN: Unblinded intervention trial with historical controls. SETTING: A university hospital, tertiary referral center for cardiothoracic surgery. PATIENTS: Consecutive patients undergoing cardiothoracic surgery between August 1, 1989, and February 1, 1991 (historical control group), and between March 1, 1991, and August 1, 1992 (intervention group). RESULTS: The historical control group consisted of 928 patients and the intervention group of 868, of whom 752 actually were treated. The 116 patients who were unintentionally not treated were considered as a concurrent control group. In the intention-to-treat analysis, a significant reduction in SSI rate was observed after the intervention (historical-control group 7.3% and intervention group 2.8%; P < .0001). The SSI rate in the concurrent control group was significantly higher than in the treated group (7.8% and 2.0%, respectively; P = .0023). Resistance of S aureus to mupirocin was not observed. CONCLUSION: The results of this study indicate that perioperative elimination of nasal carriage using mupirocin nasal ointment significantly reduces the SSI rate in cardiothoracic surgery patients and warrants a prospective, randomized, placebo-controlled efficacy trial. This preventive measure may be beneficial in other categories of surgical patients as well.

Cost-effectiveness of perioperative mupirocin nasal ointment in cardiothoracic surgery.

OBJECTIVE: To assess the cost-effectiveness of perioperative intranasal application of mupirocin calcium ointment in cardiothoracic surgery. DESIGN: Cost-effectiveness analysis based on results of an intervention study with historical controls. SETTING: University Hospital Rotterdam, a tertiary referral center for cardiac and pulmonary surgery. PATIENTS: Consecutive patients undergoing cardiothoracic surgery between August 1, 1989, and February 1, 1991 (control group, n = 928), and between March 1, 1991, and August 1, 1992 (intervention group, n = 868). INTERVENTION: Perioperative nasal application of mupirocin calcium ointment started on the day before surgery, continued for 5 days, twice daily. RESULTS: Postoperative costs were increased significantly in patients with a surgical-site infection (SSI), compared with uninfected patients (P < .001). Mean SSI-attributable costs were estimated at $16,878 (95% confidence interval, $15,575-$18,181). The incidence of SSIs was 7.3% in the control group and 2.8% in the intervention group, mupirocin effectiveness being 62%. The costs of mupirocin were $11 per patient. Thus, the savings per SSI prevented were $16,633. To validate this comparative estimate of SSI-attributable costs, a noncomparative analysis of the postoperative length of stay (POLS) was performed, according to the Appropriateness Evaluation Protocol. Approximately 50% of the comparative SSI-attributable POLS were judged SSI-attributable in the noncomparative analysis. Sensitivity analyses, testing for the robustness of our conclusions, indicated that the presented model is rather insensitive to variations in the incidence of SSIs and for the effectiveness and costs of mupirocin. SSI-attributable costs were shown to be the only variable with substantial effect on the cost-effectiveness ratio. Perioperative mupirocin would result in net costs instead of savings only if SSI-attributable costs were less than $245. CONCLUSIONS: SSIs in patients undergoing cardiothoracic surgery are associated with a substantial increase in postoperative costs. Provided that perioperative mupirocin reduces the SSI rate, this measure will be highly cost-effective in most centers providing cardiothoracic surgical services.

Bioavailability of ciprofloxacin after multiple enteral and intravenous doses in ICU patients with severe gram-negative intra-abdominal infections.

BACKGROUND: Few data are available on the pharmacokinetics of multiple enteral dosing of ciprofloxacin in critically ill intensive care patients and none for those with severe gram-negative intra-abdominal infections (GNIAI). OBJECTIVE: To determine the bioavailability of enteral ciprofloxacin in tube-fed intensive care patients with severe GNIAI. DESIGN: A randomized crossover study. SETTING: University-based medical center. PATIENTS: 5 critically ill intensive care patients with GNIAI and an estimated creatinine clearance > 25 ml/ min who received continuous tube feeding. INTERVENTIONS: Multiple doses of enteral 750 mg b.i.d. versus 400 mg b.i.d.i.v. ciprofloxacin. MEASUREMENTS: The calculated 12-h area under the serum concentration versus time curve after 750 mg b.i.d. enteral dosing was equivalent to that after 400 mg b.i.d.i.v. The mean bioavailability of enteral dosing was 53.1% [95% confidence interval (CI) 43.5-62.8]. In seven additional patients, the mean serum steady-state concentration at 2 h after enteral administration was 3.9 microg/ml (95% CI 1.9-5.9), not significantly different from that found in the crossover study (p = 0.4). CONCLUSIONS: In tube-fed intensive care patients with severe GNIAI, the bioavailability of enteral ciprofloxacin is adequate.

Genetic homogeneity among methicillin-resistant Staphylococcus aureus strains from Saudi Arabia.

Ninety-four strains of methicillin-resistant Staphylococcus aureus (MRSA) were collected from patients nursed in several hospitals in Saudi Arabia, before they were referred to King Faisal Specialist Hospital and Research Centre for tertiary care. The hospitals were from geographically diverse regions and as such the entirety of Saudi Arabia was covered. All strains were genetically typed by random amplification of polymorphic DNA (RAPD) analysis using three different primers and a representative subset of the strains was analyzed with pulsed-field gel electrophoresis (PFGE) as well. It was concluded that 87 out of 94 (93%) belong to a single clonally related lineage of MRSA. In the other 7 cases, the DNA banding patterns were shown to differ only slightly from those determined for the clonal type. PFGE analysis confirmed the homogeneity of the collection of strains. When the RAPD and PFGE fingerprints obtained for the Saudi clone were compared to those generated for a collection of MRSA with a more diverse geographical background, it was shown that the clonal type from Saudi Arabia was not identical to any of these MRSA strains. Our data provide another example of the capacity of certain MRSA clones to expand through entire nations and establish themselves permanently among large number of hospitals and, consequently, even larger numbers of patients.

Epidemiology of nosocomial fungal infections: invasive aspergillosis and the environment.

The incidence rates of invasive aspergillosis have increased dramatically during the last two decades, and, despite all diagnostic and therapeutic efforts, outcome is often fatal. Therefore, preventive measures are of major importance in the control of invasive aspergillosis, and require full understanding of the epidemiology of this devastating disease. The environment has been suggested to play a crucial role in the epidemiology of invasive aspergillosis. Aspergillus spores are released in the air and may remain airborne for prolonged periods. As a result, spores are ubiquitously found in air and contaminate anything in contact with air. It has been hypothesized that the inhalation of airborne Aspergillus spores, either directly or through intermediate nasopharyngeal colonization, is a direct cause of pulmonary infection in immunocompromised patients. Recently, water has been suggested as an additional source of "airborne" Aspergillus spp. This review summarizes the current knowledge on the role of the environment in the epidemiology of invasive aspergillosis.

Physical activity, calcium intake, and bone mineral content in children in The Netherlands.

STUDY OBJECTIVE: To examine the relation between physical activity, calcium intake, and bone mineral content in children. DESIGN: Population based, cross sectional study. SETTING: Primary schools in Zoetermeer, The Netherlands. PARTICIPANTS: Altogether 1359 Dutch boys and girls, aged 7 to 11 years (response rate 88%). MEASUREMENTS: Bone mineral content was measured by quantitative roentgen microdensitometry of the midphalanx of the second digit at the diaphyseal and metaphyseal site. Maximal exercise testing, according to the Bruce treadmill protocol, was used to assess physical fitness. Habitual physical activity was assessed by use of a questionnaire on physical activities. Daily calcium intake from dairy products was estimated by use of a semiquantitative food frequency questionnaire. MAIN RESULTS: Bone mineral content in boys was not linearly associated with physical fitness after adjustments for differences in height, body weight, chronological age, and skeletal age. In girls a linear association was found at the metaphyseal site only. When extreme groups were compared, bone mineral content was found to be higher in "high fitness children" (upper decile) than "low fitness children" (lowest decile), with statistical significance reached in boys only. When analyses were performed in subgroups of skeletal age, a clear linear relation between physical fitness and bone mineral content was seen in the mature subgroup in both boys and girls. No linear association was found between habitual physical activity and bone mineral content, while the results in extreme groups (that is, upper versus lowest decile) and in subgroups of skeletal age were comparable to those on physical fitness in boys only. No association was found between daily calcium intake and bone mineral content in this age group. CONCLUSIONS: This cross sectional study in children aged 7 to 11 years suggests that an increased bone mineral content is found only in those with a high level of physical activity. This association is most pronounced in the more mature children. No evidence was found for an association between daily calcium intake and bone mineral content in childhood.

Dutch guideline for preventing nosocomial transmission of highly resistant microorganisms (HRMO).

Hospitals are faced with the increasingly rapid emergence and dissemination of antimicrobial-resistant microorganisms. US and European guidelines on the prevention of antimicrobial resistance in hospitals were, until recently, mainly directed at methicillin-resistant Staphylococcus aureus (MRSA). In 2004, the Dutch Working Party on Infection Prevention issued a guideline on the prevention of nosocomial transmission of highly resistant microorganisms (HRMO), in order to fulfill the growing need for additional guidance on the control of other pathogens with acquired resistance and the potential to spread within hospitals (such as glycopeptide-resistant Enterococcus faecium, penicillin-resistant Streptococcus pneumoniae, extendedspectrum beta-lactamase producing Enterobacteriaceae, and other (multi)drug-resistant gram-negatives). In addition to providing criteria for defining HRMO, the Dutch guideline provides recommendations on isolation of patients, active surveillance, and contact tracing. The guideline will enable the comparison of HRMO rates between hospitals, and may be used to evaluate the efficacy of programs to control antibiotic use and/or nosocomial transmission of resistant pathogens. The eventual success of nationwide implementation of this guideline remains to be established in the coming years.

Follow-up of Staphylococcus aureus nasal carriage after 8 years: redefining the persistent carrier state.

Studies of Staphylococcus aureus nasal carriage have distinguished three carriage patterns: persistent, intermittent, and noncarriage. The criteria used to identify these carriage patterns have been inconsistent. In 1988 the S. aureus nasal carrier index, i.e., the proportion of nasal swab specimen cultures yielding S. aureus, was determined for 91 staff members of various departments of a large university hospital by obtaining weekly nasal swab specimens for culture over a 12-week period. Thirty-three (36%) persons had carrier indices of 0.80 or higher, 15 (17%) had indices between 0.1 and 0.7, and 43 (47%) had indices of zero. In 1995, 17 individuals with carrier indices of 0.80 or higher in 1988 were available for reexamination. For 12 (71%) of these individuals, S. aureus was again isolated from a single nasal swab, i.e., from each individual with a 1988 carrier index of 1.0 but from only half of those with indices below 1.0. Genotyping (by randomly amplified polymorphic DNA analysis and pulsed-field gel electrophoresis) of all S. aureus strains showed that strains isolated from only three individuals, all with 1988 carrier indices of 1.0, in 1988 and 1995 showed genetic similarity. In conclusion, persistent S. aureus nasal carriage is a unique characteristic of a fraction of the population, and the attribute "persistent" should be confined to those individuals for whom serial nasal swab specimen cultures consistently yield S. aureus.

Nasal carriage of staphylococcus aureus and epidemiology of surgical-site infections in a Sudanese university hospital.

Surgical site infections (SSI) due to Staphylococcus aureus among 256 male and 158 female patients (mean age, 28 years) undergoing elective surgery at the Soba University Hospital (Khartoum, Sudan) were studied. During an 11-month study period all patients were analyzed for nasal carriage of S. aureus at the time of admission. Follow-up of the development of SSI proceeded until 4 weeks after the operations. In addition, nasal swabs were obtained periodically during the same period from 82 members of the staff. In order to discriminate autoinfection from cross infection, bacterial isolates were typed by random amplification of polymorphic DNA (RAPD), pulsed-field gel electrophoresis (PFGE) of DNA macrorestriction fragments, and restriction fragment length polymorphism analysis of the protein A and coagulase genes. Preoperative cultures revealed the presence of S. aureus in the noses of 98 patients (24%). The overall number of postsurgical wound infections in the entire group was 57 (14%), 24 of which were due to S. aureus. Only 6 of the 98 nasal S. aureus carriers suffered from wound infections by the same species. In these six cases the infecting strain could not be genetically discriminated from the nasal inhabitant, substantiating autoinfection. However, nasal carriage of S. aureus is not a significant risk factor for the development of SSI in this setting (6 of 98 patients with autoinfection versus 18 of 316 patients [414 - 98 patients] with cross infection; P = 0.81), most probably due to the fact that noncarriers are at a significant and relatively large risk for acquiring an independent S. aureus SSI. The other S. aureus strains causing SSI showed a high degree of genetic heterogeneity, demonstrating that it is not an epidemic strain that is causing the SSI. Among the staff personnel screened, 47.4% did not carry S. aureus in the nose at any time during the study period, whereas 13. 2% persistently carried a single strain in the nose. Another 39.5% could be classified as intermittent carriers. When strains derived from staff personnel were genetically typed, it was demonstrated that most of the strains represented genetic variants clearly differing from the isolates causing SSI. On the other hand, possible cross colonization among staff personnel and even cross infection from staff personnel to patients or from patient to patient were demonstrated in some cases, but epidemic spread of a single strain or a few clonally related strains of S. aureus could be excluded.