Claire Nonhoff

Molecular Characterization of an Epidemic Clone of Panantibiotic-Resistant Pseudomonas aeruginosa

ABSTRACT We describe the molecular characterization of a multiresistant Pseudomonas aeruginosa clone causing an outbreak in the intensive care unit (ICU) of a tertiary-care university hospital. Analysis included antimicrobial susceptibility profile, O-serotyping, pulsed-field gel electrophoresis, and amplified fragment length polymorphism. Resistance mechanisms were characterized, including production of naturally occurring and acquired β-lactamases, porin expression, and efflux pump systems. Eighteen patients were colonized or infected with multiresistant P. aeruginosa. Multiresistant P. aeruginosa was panresistant to penicillins, cephalosporins, carbapenems, aminoglycosides, and fluoroquinolones and remained susceptible only to colistin. Sixteen isolates (89%) belonged to serotype O:11, pulsed-field gel electrophoresis type A1, and amplified fragment length polymorphism type A. Resistance characterization of this epidemic clone showed an overexpression of the chromosomal cephalosporinase AmpC combined with decreased expression of porin OprD and the absence of metallo-β-lactamase or extended-spectrum beta-lactamase. An upregulation of the MexXY efflux system due to an agrZ mutation in the mexZ repressor was detected. This epidemic clone was restricted to the ICU and was not found elsewhere in hospital. Contamination of the ICU environment and the hands of an ICU nurse with this clone suggests possible hand-borne transmission. Implementation of contact precautions effectively controlled transmission of the epidemic clone. This study illustrates the ability of multiresistant P. aeruginosa to cause an outbreak with significant morbidity and mortality and underscores the need to identify clonal outbreaks, which require targeted infection control measures.

Controlled Evaluation of the IDI-MRSA Assay for Detection of Colonization by Methicillin-Resistant Staphylococcus aureus in Diverse Mucocutaneous Specimens

ABSTRACT Rapid and reliable detection of methicillin-resistant Staphylococcus aureus (MRSA) carriers is crucial for the effective control of MRSA transmission in healthcare facilities. The aim of this study was to verify the performance of the IDI-MRSA real-time PCR assay for direct MRSA detection in diverse mucocutaneous swabs from hospitalized patients. Swabs from nares (n = 522) and skin or other superficial sites (n = 478) were prospectively collected for MRSA screening from 466 patients admitted to an 858-bed teaching hospital. Swabs were inoculated onto selective chromogenic MRSA-ID agar, buffer extraction solution for IDI-MRSA assay, and enrichment broth. MRSA was detected by culture in 100 specimens from 47 patients. Compared to enrichment culture, the sensitivity and specificity of the PCR assay were 81.0 and 97.0%, respectively, and its positive and negative predictive values were 75.0 and 97.9%, respectively. The IDI-MRSA assay was more sensitive on swabs from nares (90.6%) than from other body sites (76.5%, P < 0.01). The PCR assay detected MRSA in 42 of 47 patients with culture positive study samples. Of 26 patients with culture-negative but PCR-positive study samples, 11 were probable true MRSA carriers based on patient history and/or positive culture on a new sample. The median turnaround time for PCR results was 19 h versus 3 days for agar culture results and 6 days for enrichment culture results. These data confirm the value of IDI-MRSA assay for rapid screening of MRSA mucocutaneous carriage among hospitalized patients. Cost-effectiveness studies are warranted to evaluate the impact of this assay on infection control procedures in healthcare settings.

National Epidemiologic Surveys of Enterobacter aerogenes in Belgian Hospitals from 1996 to 1998

ABSTRACT Two national surveys were conducted to describe the incidence and prevalence of Enterobacter aerogenes in 21 Belgian hospitals in 1996 and 1997 and to characterize the genotypic diversity and the antimicrobial resistance profiles of clinical strains ofE. aerogenes isolated from hospitalized patients in Belgium in 1997 and 1998. Twenty-nine hospitals collected 10 isolates ofE. aerogenes, which were typed by arbitrarily primed PCR (AP-PCR) using two primers and pulsed-field gel electrophoresis. MICs of 10 antimicrobial agents were determined by the agar dilution method. Beta-lactamases were detected by the double-disk diffusion test and characterized by isoelectric point. The median incidence of E. aerogenes colonization or infection increased from 3.3 per 1,000 admissions in 1996 to 4.2 per 1000 admissions in the first half of 1997 (P < 0.01). E. aerogenes strains (n = 260) clustered in 25 AP-PCR types. Two major types, BE1 and BE2, included 36 and 38% of strains and were found in 21 and 25 hospitals, respectively. The BE1 type was indistinguishable from a previously described epidemic strain in France. Half of the strains produced an extended-spectrum beta-lactamase, either TEM-24 (in 86% of the strains) or TEM-3 (in 14% of the strains). Over 75% of the isolates were resistant to ceftazidime, piperacillin-tazobactam, and ciprofloxacin. Over 90% of the strains were susceptible to cefepime, carbapenems, and aminoglycosides. In conclusion, these data suggest a nationwide dissemination of two epidemic multiresistantE. aerogenes strains in Belgian hospitals. TEM-24 beta-lactamase was frequently harbored by one of these epidemic strains, which appeared to be genotypically related to a TEM-24-producing epidemic strain from France, suggesting international dissemination.

In Vitro Activities of Ceftobiprole, Tigecycline, Daptomycin, and 19 Other Antimicrobials against Methicillin-Resistant Staphylococcus aureus Strains from a National Survey of Belgian Hospitals

ABSTRACT The in vitro activities of 22 antimicrobial agents, including ceftobiprole, daptomycin, and tigecycline, against 511 methicillin-resistant Staphylococcus aureus (MRSA) isolates from 112 Belgian hospitals were studied by using the CLSI agar dilution method. Isolates were characterized by pulsed-field gel electrophoresis (PFGE) analysis and by PCR detection of determinants of resistance to aminoglycosides, macrolides-lincosamides-streptogramins, and tetracyclines. A representative set of isolates with different PFGE genotypes was further characterized by multilocus sequence typing, determination of staphylococcal cassette chromosome mec (SCCmec) type, and multiplex PCR for toxic shock syndrome type 1 (TSST-1) and Panton-Valentine leukocidin genes. MRSA isolates belonged to nine epidemic MRSA clones, of which sequence type 45 (ST45)-SCCmec IV and ST8-SCCmec IV were predominant, accounting for 49 and 20% of isolates, respectively. The distribution of antimicrobial resistance and TSST-1 genes was strongly linked to clonal types. Ceftobiprole, daptomycin, and tigecycline showed high activity against all isolates of these sporadic and epidemic MRSA clones, as indicated by MIC90s of 2 mg/liter, 0.5 mg/liter, and 0.25 mg/liter, respectively. The MIC distribution of daptomycin and tigecycline was not different in isolates with decreased susceptibility to glycopeptides or tetracyclines, respectively. Ceftobiprole MICs were not correlated with oxacillin and cefoxitin MICs. These data indicate excellent activity of the newly developed agents ceftobiprole, daptomycin, and tigecycline against MRSA isolates recently recovered from hospitalized patients in Belgium, supporting their therapeutic potential for nosocomial MRSA infections.

Epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) among residents of nursing homes in Belgium

OBJECTIVES A national survey was conducted to determine the prevalence, risk factors and molecular epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) carriage among nursing home (NH) residents in Belgium. METHODS A random stratified, cross-sectional prevalence survey was conducted in NH residents who were screened for MRSA carriage by multisite enriched culture. Characteristics of NHs and residents were collected by a questionnaire survey and analysed by two-stage logistic regression modelling. MRSA isolates were genotyped by PFGE, staphylococcal cassette chromosome mec (SCCmec) typing, multilocus sequence typing (MLST) and resistance genes. RESULTS Of 2953 residents screened in 60 NHs, 587 (19.9%) were MRSA carriers. Risk factors included hospital contact, antibiotic exposure, impaired mobility and skin lesions at the resident level, and lack of MRSA surveillance, lack of antibiotic therapeutic formulary and the combination of less-developed infection control activities and a high ratio of physicians to residents at the institution level. MRSA isolates showed eight major types, three of which were predominant: B2-ST45-SCCmec IV (49%; where ST stands for sequence type); A21-ST8-SCCmec IV (13%); and A20-ST8-SCCmec IV (10%). Each was recovered in 55, 21 and 25 NHs, respectively. The geographical distribution of NH genotypes paralleled that of acute-care hospitals. CONCLUSIONS A high prevalence of MRSA carriage in NH residents was associated with hospital care, co-morbidities and less-developed coordination of institutional care. The predominant MRSA strains from NH residents and hospitalized patients of the same area were identical. Strengthening and coordination of MRSA surveillance and control activities are warranted within and between NHs and hospitals.

National Surveillance of Methicillin-Resistant Staphylococcus aureus in Belgian Hospitals Indicates Rapid Diversification of Epidemic Clones

ABSTRACT Methicillin-resistant Staphylococcus aureus (MRSA) strains (n = 455) collected in 2001 from 100 Belgian hospitals were characterized by molecular typing and by resistance gene distribution to macrolides-lincosamides-streptogramins and to aminoglycoside antibiotics. Rapid diversification of MRSA clones, compared with results of previous surveys, was evidenced by the broad geographic distribution of seven major clones belonging to the pandemic MRSA clonal complexes 5, 8, 22, 30, and 45 by multilocus sequence typing.

Comparative Epidemiology of Staphylococcus epidermidis Isolates from Patients with Catheter-Related Bacteremia and from Healthy Volunteers

ABSTRACT Staphylococcus epidermidis is a major cause of catheter-related bloodstream infections (CRBSIs). Recent studies suggested the existence of well-adapted, highly resistant, hospital-associated S. epidermidis clones. The molecular epidemiology of S. epidermidis in Belgian hospitals and the Belgian community has not been explored yet. We compared a set of 33 S. epidermidis isolates causing CRBSI in hospitalized patients with a set of 33 commensal S. epidermidis isolates. The factors analyzed included resistance to antibiotics and genetic diversity as determined by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), and SCCmec typing. Additionally, the presence of virulence-associated mobile genetic elements, the ica operon and the arginine catabolic mobile element (ACME), was assessed and compared against clinical data. CRBSI S. epidermidis isolates were significantly resistant to more antibiotics than commensal S. epidermidis isolates. The two populations studied were very diverse and genetically distinct as only 23% of the 37 PFGE types observed were harbored by both CRBSI and commensal isolates. ACME was found in 76% of S. epidermidis strains, regardless of their origin, while the ica operon was significantly more prevalent in CRBSI isolates than in commensal isolates (P < 0.05). Nine patients presented a clinically severe CRBSI, eight cases of which were due to an ica-positive multiresistant isolate belonging to sequence type 2 (ST2) or ST54. S. epidermidis isolates causing CRBSI were more resistant and more often ica positive than commensal S. epidermidis isolates, which were genetically heterogeneous and susceptible to the majority of antibiotics tested. Clinically severe CRBSIs were due to isolates belonging to two closely related MLST types, ST2 and ST54.

Performance of the Verigene Gram-Negative Blood Culture Assay for Rapid Detection of Bacteria and Resistance Determinants

ABSTRACT Nonduplicate blood cultures that were positive for Gram-negative bacilli (n = 125) were tested by the Verigene Gram-negative blood culture (BC-GN) assay; 117 (90.7%) isolates were members of the panel. For identification and resistance markers, the agreements with routine methods were 97.4% (114/117) and 92.3% (12/13). The BC-GN assay is a rapid and accurate tool for the detection of pathogens from blood cultures and could be integrated alongside conventional systems to enable faster patient management, but the clinical benefits should be further evaluated.

Epidemiology of multidrug-resistant microorganisms among nursing home residents in Belgium.

Objectives A national survey was conducted to determine the prevalence and risk factors of methicillin-resistant Staphylococcus aureus (MRSA), extended-spectrum β-lactamases-producing Enterobacteriaceae (ESBLE) and vancomycin-resistant enterococci (VRE) among nursing home residents in Belgium. Methods A random stratified, national prevalence survey was conducted in nursing home residents who were screened for carriage of ESBLE, MRSA and VRE by multisite enriched culture. Characteristics of nursing homes and residents were collected by a questionnaire survey and were analysed by multilevel logistic regression analysis. Results Of 2791 screened residents in 60 participating nursing home, the weighted prevalence of ESBLE and MRSA carriage were 6.2% (range: 0 to 20%) and 12.2% (range: 0 to 36%), respectively. No cases of VRE were found. No relationship was found between ESBLE and MRSA prevalence rates within nursing homes and the rate of co-colonization was very low (0.8%). Geographical variations in prevalence of MRSA and ESBLE and in distribution of ESBL types in nursing home residents paralleled that of acute hospitals. Risk factors of ESBLE carriage included previously known ESBLE carriage, male gender, a low level of mobility and previous antibiotic exposure. Risk factors for MRSA colonization were: previously known MRSA carriage, skin lesions, a low functional status and antacid use. Conclusions A low prevalence of ESBLE carriage was found in nursing home residents in Belgium. The prevalence of MRSA carriage decreased substantially in comparison to a similar survey conducted in 2005. A low functional status appeared as a common factor for ESBLE and MRSA carriage. Previous exposure to antibiotics was a strong predictor of ESBLE colonization while increased clustering of MRSA carriage suggested the importance of cross-transmission within nursing homes for this organism. These results emphasize the need for global coordination of the surveillance of MDRO within and between nursing homes and hospitals.

Emergence and spread of gentamicin-susceptible strains of methicillin-resistant Staphylococcus aureus in Belgian hospitals

The aim of this study was to follow the evolution of the clonal distribution and antimicrobial susceptibility of clinical strains of methicillin-resistant Staphylococcus aureus (MRSA) recovered from Belgian hospitals between 1995 and 1997-1998. MRSA strains were genotyped by inter-IS256 spacer length polymorphism PCR and SmaI macrorestriction analysis. MICs of 18 antimicrobials were determined by the agar dilution method. MRSA strains from the 1997-1998 survey were further tested by vancomycin screen agar, E-test, broth microdilution methods, and population analysis. Between 1995 and 1997-1998, epidemic group A strains decreased in proportion from 73% to 44%, whereas MRSA Group B and C strains increased from 17% to 38% and from 5% to 8%. The proportion of strains susceptible to gentamicin increased between the surveys from 22% to 48%. This was associated with a higher proportion of group B and C strains in the last survey. Heterogeneous vancomycin-intermediate S. aureus (hetero-VISA) strains were found in 2% isolates from 1997 to 1998. These hetero-VISA isolates were genotypically related to the MRSA group A strains and were resistant to gentamicin. In conclusion, two emerging epidemic MRSA genotypes, susceptible to gentamicin, have spread among Belgian hospitals during the 1990s. Hetero-VISA were present at low frequency among MRSA strains belonging to a widespread endemic genotype.