Silke Peter

cyp51A-Based Mechanisms of Aspergillus fumigatus Azole Drug Resistance Present in Clinical Samples from Germany

ABSTRACT Since the mid-1990s, a steady increase in the occurrence of itraconazole-resistant Aspergillus fumigatus isolates has been observed in clinical contexts, leading to therapeutic failure in the treatment of aspergillosis. This increase has been predominantly linked to a single allele of the cyp51A gene, termed TR/L98H, which is thought to have arisen through the use of agricultural azoles. Here, we investigated the current epidemiology of triazole-resistant A. fumigatus and underlying cyp51A mutations in clinical samples in Germany. From a total of 527 samples, 17 (3.2%) showed elevated MIC0 values (the lowest concentrations with no visible growth) for at least one of the three substances (itraconazole, voriconazole, and posaconazole) tested. The highest prevalence of resistant isolates was observed in cystic fibrosis patients (5.2%). Among resistant isolates, the TR/L98H mutation in cyp51A was the most prevalent, but isolates with the G54W and M220I substitutions and the novel F219C substitution were also found. The isolate with the G54W substitution was highly resistant to both itraconazole and posaconazole, while all others showed high-level resistance only to itraconazole. For the remaining six isolates, no mutations in cyp51A were found, indicating the presence of other mechanisms. With the exception of the strains carrying the F219C and M220I substitutions, many itraconazole-resistant strains also showed cross-resistance to voriconazole and posaconazole with moderately increased MIC0 values. In conclusion, the prevalence of azole-resistant A. fumigatus in our clinical test set is lower than that previously reported for other countries. Although the TR/L98H mutation frequently occurs among triazole-resistant strains in Germany, it is not the only resistance mechanism present.

A multicentre cohort study on colonization and infection with ESBL-producing Enterobacteriaceae in high-risk patients with haematological malignancies

BACKGROUND Bloodstream infections (BSIs) caused by enterobacteria remain a leading cause of mortality in patients with chemotherapy-induced neutropenia. The rate and type of colonization and infection with ESBL-producing Enterobacteriaceae (ESBL-E) and their mode of transmission in German cancer centres is largely unknown. METHODS We performed a prospective, observational study at five German university-based haematology departments. Participating sites screened for intestinal ESBL-E colonization within 72 h of admission, every 10 ± 2 days thereafter and before discharge. Three of the five centres performed contact isolation for patients colonized or infected with ESBL-E. Molecular characterization of resistance mechanisms and epidemiological typing of isolates by repetitive extragenic palindromic PCR (rep-PCR) and PFGE was performed to assess strain transmission between patients. RESULTS Between October 2011 and December 2012, 719 hospitalizations of 497 haematological high-risk patients comprising 20,143 patient-days were analysed. Mean duration of in-hospital stay was 36.6 days (range: 2-159 days). ESBL-E were identified from screening samples (82.8% Escherichia coli and 14.6% Klebsiella pneumoniae) in 55/497 patients (11.1%; range by centre: 5.8%-23.1%). PFGE and rep-PCR revealed only a single case of potential cross-transmission among two patients colonized with K. pneumoniae. Six episodes of BSI with ESBL-E were observed. Multivariate analysis revealed previous colonization with ESBL-E as the most important risk factor for BSI with ESBL-E (OR 52.00; 95% CI 5.71-473.89). CONCLUSIONS Even though BSI with ESBL-E is still rare in this high-risk population, colonization rates are substantial and vary considerably between centres. In-hospital transmission of ESBL-E as assessed by molecular typing was the exception.

Colonization with third-generation cephalosporin-resistant Enterobacteriaceae on hospital admission: prevalence and risk factors

OBJECTIVES The objectives of this study were to prospectively assess the rectal carriage rate of third-generation cephalosporin-resistant Enterobacteriaceae (3GCREB) in non-ICU patients on hospital admission and to investigate resistance mechanisms and risk factors for carriage. METHODS Adult patients were screened for 3GCREB carriage at six German tertiary care hospitals in 2014 using rectal swabs or stool samples. 3GCREB isolates were characterized by phenotypic and molecular methods. Each patient answered a questionnaire about potential risk factors for colonization with MDR organisms (MDROs). Univariable and multivariable risk factor analyses were performed to identify factors associated with 3GCREB carriage. RESULTS Of 4376 patients, 416 (9.5%) were 3GCREB carriers. Escherichia coli was the predominant species (79.1%). ESBLs of the CTX-M-1 group (67.3%) and the CTX-M-9 group (16.8%) were the most frequent β-lactamases. Five patients (0.11%) were colonized with carbapenemase-producing Enterobacteriaceae. The following risk factors were significantly associated with 3GCREB colonization in the multivariable analysis (P < 0.05): centre; previous MDRO colonization (OR = 2.12); antibiotic use within the previous 6 months (OR = 2.09); travel outside Europe (OR = 2.24); stay in a long-term care facility (OR = 1.33); and treatment of gastroesophageal reflux disease (GERD) (OR = 1.22). CONCLUSIONS To our knowledge, this is the largest admission prevalence study of 3GCREB in Europe. The observed prevalence of 9.5% 3GCREB carriage was higher than previously reported and differed significantly among centres. In addition to previously identified risk factors, the treatment of GERD proved to be an independent risk factor for 3GCREB colonization.

Analysis of a long-term outbreak of XDR Pseudomonas aeruginosa: a molecular epidemiological study

OBJECTIVES Here we report on a long-term outbreak from 2009 to 2012 with an XDR Pseudomonas aeruginosa on two wards at a university hospital in southern Germany. METHODS Whole-genome sequencing was performed on the outbreak isolates and a core genome was constructed for molecular epidemiological analysis. We applied a time-place-sequence algorithm to improve estimation of transmission probabilities. RESULTS By using conventional infection control methods we identified 49 P. aeruginosa strains, including eight environmental isolates that belonged to ST308 (by MLST) and carried the metallo-β-lactamase IMP-8. Phylogenetic analysis on the basis of a non-recombinant core genome that contained 22 outbreak-specific SNPs revealed a pattern of four dominant clades with a strong phylogeographic structure and allowed us to determine the potential temporal origin of the outbreak to July 2008, 1 year before the index case was diagnosed. Superspreaders at the root of clades exhibited a high number of probable and predicted transmissions, indicating their exceptional position in the outbreak. CONCLUSIONS Our results suggest that the initial expansion of dominant sublineages was driven by a few superspreaders, while environmental contamination seemed to sustain the outbreak for a long period despite regular environmental control measures.

Evaluation of the Accelerate Pheno System for Fast Identification and Antimicrobial Susceptibility Testing from Positive Blood Cultures in Bloodstream Infections Caused by Gram-Negative Pathogens

ABSTRACT Bloodstream infections (BSI) are an important cause of morbidity and mortality. Increasing rates of antimicrobial-resistant pathogens limit treatment options, prompting an empirical use of broad-range antibiotics. Fast and reliable diagnostic tools are needed to provide adequate therapy in a timely manner and to enable a de-escalation of treatment. The Accelerate Pheno system (Accelerate Diagnostics, USA) is a fully automated test system that performs both identification and antimicrobial susceptibility testing (AST) directly from positive blood cultures within approximately 7 h. In total, 115 episodes of BSI with Gram-negative bacteria were included in our study and compared to conventional culture-based methods. The Accelerate Pheno system correctly identified 88.7% (102 of 115) of all BSI episodes and 97.1% (102 of 105) of isolates that are covered by the systems identification panel. The Accelerate Pheno system generated an AST result for 91.3% (95 of 104) samples in which the Accelerate Pheno system identified a Gram-negative pathogen. The overall category agreement between the Accelerate Pheno system and culture-based AST was 96.4%, the rates for minor discrepancies 1.4%, major discrepancies 2.3%, and very major discrepancies 1.0%. Of note, ceftriaxone, piperacillin-tazobactam, and carbapenem resistance was correctly detected in blood culture specimens with extended-spectrum beta-lactamase-producing Escherichia coli (n = 7) and multidrug-resistant Pseudomonas aeruginosa (n = 3) strains. The utilization of the Accelerate Pheno system reduced the time to result for identification by 27.49 h (P < 0.0001) and for AST by 40.39 h (P < 0.0001) compared to culture-based methods in our laboratory setting. In conclusion, the Accelerate Pheno system provided fast, reliable results while significantly improving turnaround time in blood culture diagnostics of Gram-negative BSI.

Prevalence and risk factors associated with colonization and infection of extensively drug-resistant Pseudomonas aeruginosa: a systematic review

Pseudomonas aeruginosa is a Gram-negative human pathogen with extensively drug-resistant (XDR) strains emerging in hospitals across the globe. This systematic review is focused on the worldwide prevalence of XDR P. aeruginosa (XDR-PA) and on the risk factors associated with its colonization and infection, based on literature available through PubMed, Web of Science and BioMed Central databases. An overview of surveillance systems is provided as well as a synopsis on the prevalence of XDR-PA, showing an increase in recent reports. Risk factors independently associated with XDR-PA colonization or infections are described in four groups with reference to antimicrobial therapy, medical devices as well as patient- and hospital environment-related factors.

Effect of metallo-β-lactamase production and multidrug resistance on clinical outcomes in patients with Pseudomonas aeruginosa bloodstream infection: a retrospective cohort study

BackgroundBlood stream infections (BSI) with Pseudomonas aeruginosa lead to poor clinical outcomes. The worldwide emergence and spread of metallo-β-lactamase (MBL) producing, often multidrug-resistant organisms may further aggravate this problem. Our study aimed to investigate the effect of MBL-producing P. aeruginosa (MBL-PA) and various other resistance phenotypes on clinical outcomes.MethodsA retrospective cohort study was conducted in three German hospitals. Medical files from 2006 until 2012 were studied, and a number of 113 patients with P. aeruginosa BSI were included. The presence of VIM, IMP and NDM genes was detected using molecular techniques. Genetic relatedness was assessed through multilocus sequence typing (MLST). The effect of resistance patterns or MBL production on clinical outcomes was investigated by using multivariate Cox regression models.ResultsIn-hospital mortality was significantly higher in patients with MBL-PA and multidrug-resistant P. aeruginosa. However, neither BSI with MBL-PA nor BSI with various resistance phenotypes of P. aeruginosa were independently associated with mortality or length of hospital stay. In multivariate models, the SAPS II score (HR 1.046), appropriate definitive treatment (HR range 0.25-0.26), and cardiovascular disease (HR range 0.44-0.46) were independent predictors of mortality. Concomitant infections were associated with an excess length of stay (HR < 1).ConclusionsMedication with appropriate antimicrobial agents at any time during the course of infection remains the key for improving clinical outcomes in patients with P. aeruginosa BSI and should be combined with a strict implementation of routine infection control measures.

Time series analysis as a tool to predict the impact of antimicrobial restriction in antibiotic stewardship programs using the example of multidrug-resistant Pseudomonas aeruginosa

ABSTRACT The association between antimicrobial consumption and resistance in nonfermentative Gram-negative bacteria is well-known. Antimicrobial restriction, implemented in clinical routines by antibiotic stewardship programs (ASPs), is considered a means to reduce resistance rates. Whether and how antimicrobial restriction can accomplish this goal is still unknown though. This leads to an element of uncertainty when designing strategies for ASPs. From January 2002 until December 2011, an observational study was performed at the University Hospital Tübingen, Tübingen, Germany, to investigate the association between antimicrobial use and resistance rates in Pseudomonas aeruginosa. Transfer function models were used to determine such associations and to simulate antimicrobial restriction strategies. Various positive associations between antimicrobial consumption and resistance were observed in our setting. Surprisingly, impact estimations of different antimicrobial restriction strategies revealed relatively low intervention expenses to effectively attenuate the observed increase in resistance. For example, a simulated intervention of an annual 4% reduction in the use of meropenem over 3 years from 2009 until 2011 yielded a 62.5% attenuation (95% confidence interval, 15% to 110%) in the rising trend of multidrug-resistant Pseudomonas aeruginosa (three- and four-class-resistant P. aeruginosa [34MRGN-PA]). Time series analysis models derived from past data may be a tool to predict the outcome of antimicrobial restriction strategies, and could be used to design ASPs.

Antibiotic susceptibility of Clostridium difficile is similar worldwide over two decades despite widespread use of broad-spectrum antibiotics: an analysis done at the University Hospital of Zurich

BackgroundClostridium difficile infection (CDI) remains a major health problem worldwide. Antibiotic use, in general, and clindamycin and ciprofloxacin, in particular, have been implicated in the pathogenesis of CDI. Here, we hypothesized that antibiotics that are highly active in vitro against C. difficile are less frequently associated with CDI than others. The primary goals of our study were to determine if antibiotic susceptibility and CDI are associated and whether the antimicrobial susceptibility of C. difficile changed over the years.Methods and resultsWe examined a large panel of C. difficile strains collected in 2006-2008 at the University Hospital of Zurich. We found that the antimicrobial susceptibilities to amoxicillin/clavulanate, piperacillin/tazobactam, meropenem, clindamycin, ciprofloxacin, ceftriaxone, metronidazole and vancomycin were similar to those reported in the literature and that they are similar to those reported in other populations over the last two decades. Antibiotic activity did not prevent CDI. For example, thre use of meropenem, which is highly active against all strains tested, was a clear risk factor for CDI. Most of the antibiotics tested also showed a higher minimum inhibitory concentration distribution than that of EUCAST. All strains were susceptible to metronidazole. One strain was resistant to vancomycin.ConclusionsAntibiotic susceptibilities of the collection of C. difficile from the University Hospital of Zurich are similar to those reported by others since the 1980. Patients treated with carbapenems and cephalosporins had the highest risk of developing CDI irrespective of the antimicrobial activity of carbapenems.

Time to positivity as prognostic tool in patients with Pseudomonas aeruginosa bloodstream infection

OBJECTIVES The time to positivity (TTP), measured as the time span between the start of incubation and the alert signal from the blood culture device, has been described as useful tool of prognosis in patients suffering from blood stream infection with Staphylococcus aureus, Escherichia coli and Klebsiella pneumonia. The present study investigates the relationship between TTP and in-hospital mortality in patients with monomicrobial Pseudomonas aeruginosa blood stream infection (PA-BSI). METHODS From 2006 until 2012 a retrospective cohort study was undertaken in 3 hospitals in the region surrounding Tübingen, Germany. Seventy-four patients with monomicrobial PA-BSI were studied. TTP and clinical parameters were determined and analyzed by receiver operating characteristic (ROC) analysis and Cox regression. RESULTS The in-hospital mortality of our clinical cohort was 33.78%. In multivariate Cox regression, a TTP ≤ 18 h proved to be independently associated with mortality (HR 3.83, P = 0.012) along with SAPS II score (HR 1.04, P = 0.006), cardiac disease (HR 0.33, P = 0.008) and appropriate definitive antimicrobial treatment (HR 0.21, P = 0.013). CONCLUSIONS TTP is an easy-to-measure laboratory tool for prognosis in patients with monomicrobial PA-BSI, providing useful information in addition to clinical parameters.