Silke Polsfuss

University of Zurich, Zürich, ZH, Switzerland

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Publications (10)35.15 Total impact

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    ABSTRACT: This study aimed at evaluating the AID ESBL line probe assay for the detection of extended-spectrum β-lactamase (ESBL) and KPC carbapenemase genes in Enterobacteriaceae. The AID ESBL line probe assay was verified for accuracy of its probes using PCR products from clinical ESBL Enterobacteriaceae strains harbouring TEM, SHV and CTX-M ESBL genes and KPC genes and mutant fusion PCR products generated from Enterobacteriaceae strains containing wild-type (wt) TEM and wt SHV. Sensitivity and specificity was determined testing a set of 424 clinical Enterobacteriaceae strains (including 170 strains negative for TEM, SHV, CTX-M and KPC to evaluate the possibility of false positive signals). The line probe assay was shown to detect with 100% accuracy ESBL genes for which oligonucleotide probes are present in the assay. Testing a set of 424 clinical Enterobacteriaceae strains showed 100% sensitivity and specificity for the detection and differentiation of TEM, SHV and CTX-M ESBL genes present in that group. In addition, the line probe assay detected KPC genes accurately. The AID ESBL line probe assay is an accurate and easy-to-use test for the detection of ESBL and KPC genes, which can readily be implemented in the diagnostic laboratory.
    Journal of Antimicrobial Chemotherapy 09/2013; · 5.34 Impact Factor
  • 84. Jahresversammlung der Deutschen Gesellschaft für Hals-Nasen-Ohren-Heilkunde, Kopf- und Hals-Chirurgie; 05/2013
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    ABSTRACT: To compare the performance of European Committee on Antimicrobial Susceptibility Testing (EUCAST) and CLSI breakpoints following their revision in 2010, for the detection of extended-spectrum β-lactamase (ESBL) production in Enterobacteriaceae. 236 well-characterized clinical isolates (including 118 ESBL producers) were investigated by antibiotic disc testing with cefpodoxime, ceftriaxone, cefepime, cefotaxime EUCAST (5 μg/disc), ceftazidime EUCAST (10 μg/disc), cefotaxime CLSI (30 μg/disc) and ceftazidime CLSI (30 μg/disc) with the Kirby-Bauer method. Additionally, synergy phenomena were recorded between amoxicillin/clavulanic acid discs (20/10 μg/disc) and cefepime (30 μg/disc), EUCAST cefotaxime (5 μg/disc), EUCAST ceftazidime (10 μg/disc), CLSI cefotaxime (30 μg/disc) and CLSI ceftazidime [30 μg/disc; disc approximation method (DAM)]. Overall sensitivity of the cefotaxime EUCAST non-susceptible breakpoint equalled sensitivity of the cefotaxime CLSI ESBL screening breakpoint (99.2%). With the ceftazidime EUCAST non-susceptible breakpoint, 27/118 ESBL-producing isolates were not detected, whereas the ceftazidime CLSI ESBL screening breakpoint missed 41/118 ESBL-producing isolates. For cefpodoxime the resistant EUCAST breakpoint showed higher sensitivity for ESBL detection compared with the CLSI ESBL screening breakpoint/disc content (100% versus 98.3%, respectively). Sensitivities of ceftazidime and cefotaxime DAM with CLSI or EUCAST disc contents were comparable (sensitivities ranging from 84.7% to 89.8%). DAM with cefepime displayed the highest overall sensitivity (96.6%). In AmpC-producing isolates, synergy of amoxicillin/clavulanic acid with cefepime showed sensitivity and specificity for ESBL detection of 100% and 97.4%, respectively. EUCAST non-susceptible breakpoints for ceftazidime and cefpodoxime detect more ESBL-producing Enterobacteriaceae isolates compared with corresponding CLSI ESBL screening breakpoints. Implementation of the cefepime DAM can facilitate ESBL screening, especially in strains producing an AmpC β-lactamase since the test shows high sensitivity and specificity.
    Journal of Antimicrobial Chemotherapy 01/2012; 67(1):159-66. · 5.34 Impact Factor
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    ABSTRACT: Clin Microbiol Infect ABSTRACT: This study aimed to develop a modular, diagnostic algorithm for extended spectrum β-lactamase (ESBL) detection in Enterobacteriaceae. Clinical Enterobacteriaceae strains (n = 2518) were screened for ESBL production using Clinical and Laboratory Standards Institute (CLSI) breakpoints for third-generation cephalosporins and by synergy image detection (clavulanic acid/extended-spectrum cephalosporins). Isolates screening positive for ESBL (n = 242, 108 by critical CLSI diameters alone, five by double disk synergy test (DDST) alone, and 129 by both critical diameters and DDST) and 138 ESBL screening negative isolates (control group) were investigated by molecular methods considered to be the reference standard (multiplex CTX-M type PCR, TEM and SHV type sequence characterization). One hundred and twenty-four out of 242 Enterobacteriaceae isolates screening positive for ESBL were confirmed to be ESBL positive by the reference standard, the majority of them in E. coli, K. pneumoniae and E. cloacae (94, 17 and nine isolates, respectively). Prevalence of ESBL production ranged from <1% for P. mirabilis to 4.7%, 5.1% and 6.6%, for K. pneumoniae, E. cloacae and E. coli, respectively. Combining CLSI ceftriaxone and cefpodoxime critical ESBL diameters was found to be the most sensitive phenotypic screening method (sensitivity 99.2%). Combining critical diameters of cefpodoxime and ceftriaxone with DDST for cefpodoxime resulted in a sensitivity of 100%. For phenotypic confirmation, combining the CLSI recommended combined disk test (CDT) for ceftazidime and cefotaxime amended with a cefepime CDT was highly sensitive (100%) and specific (97.5%). With respect to the studied population, the diagnostic ESBL algorithm developed would have resulted in sensitivity and specificity of 100%. The corresponding flow chart is simple, easy to use, inexpensive and applicable in the routine diagnostic laboratory.
    Clinical Microbiology and Infection 11/2011; · 4.58 Impact Factor
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    ABSTRACT: Clin Microbiol Infect ABSTRACT: This study aimed to develop a modular, diagnostic algorithm for extended spectrum β-lactamase (ESBL) detection in Enterobacteriaceae. Clinical Enterobacteriaceae strains (n = 2518) were screened for ESBL production using Clinical and Laboratory Standards Institute (CLSI) breakpoints for third-generation cephalosporins and by synergy image detection (clavulanic acid/extended-spectrum cephalosporins). Isolates screening positive for ESBL (n = 242, 108 by critical CLSI diameters alone, five by double disk synergy test (DDST) alone, and 129 by both critical diameters and DDST) and 138 ESBL screening negative isolates (control group) were investigated by molecular methods considered to be the reference standard (multiplex CTX-M type PCR, TEM and SHV type sequence characterization). One hundred and twenty-four out of 242 Enterobacteriaceae isolates screening positive for ESBL were confirmed to be ESBL positive by the reference standard, the majority of them in E. coli, K. pneumoniae and E. cloacae (94, 17 and nine isolates, respectively). Prevalence of ESBL production ranged from <1% for P. mirabilis to 4.7%, 5.1% and 6.6%, for K. pneumoniae, E. cloacae and E. coli, respectively. Combining CLSI ceftriaxone and cefpodoxime critical ESBL diameters was found to be the most sensitive phenotypic screening method (sensitivity 99.2%). Combining critical diameters of cefpodoxime and ceftriaxone with DDST for cefpodoxime resulted in a sensitivity of 100%. For phenotypic confirmation, combining the CLSI recommended combined disk test (CDT) for ceftazidime and cefotaxime amended with a cefepime CDT was highly sensitive (100%) and specific (97.5%). With respect to the studied population, the diagnostic ESBL algorithm developed would have resulted in sensitivity and specificity of 100%. The corresponding flow chart is simple, easy to use, inexpensive and applicable in the routine diagnostic laboratory.
    Clinical Microbiology and Infection 11/2011; · 4.58 Impact Factor
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    ABSTRACT: Two mechanisms account for AmpC activity in Escherichia coli, namely, mutations in the ampC promoter and attenuator regions resulting in ampC overexpression and acquisition of plasmid-carried ampC genes. In this study, we analyzed 51 clinical E. coli isolates with reduced susceptibility to amoxicillin-clavulanic acid, piperacillin-tazobactam, or extended-spectrum cephalosporins for the presence of AmpC production. Three phenotypic AmpC confirmation assays (cefoxitin-cloxacillin disk diffusion test, cefoxitin-EDTA disk diffusion test, and AmpC Etest) were compared for the detection of AmpC activity. All 51 isolates were characterized genetically by mutational analysis of the chromosomal ampC promoter/attenuator region and by PCR detection of plasmid-carried ampC genes. Altogether, 21/51 (41%) E. coli isolates were considered true AmpC producers. AmpC activity due to chromosomal ampC promoter/attenuator mutations was found in 12/21 strains, and plasmid-carried ampC genes were detected in 8/21 isolates. One strain contained both ampC promoter mutations and a plasmid-carried ampC gene. All three phenotypic tests were able to detect the majority (>90%) of AmpC-positive strains correctly. Cefoxitin resistance was found to be a discriminative parameter, detecting 20/21 AmpC-producing strains. Susceptibility to extended-spectrum cephalosporins, e.g., ceftriaxone, ceftazidime, and cefotaxime, was found in 9 of the 21 AmpC-positive strains. Considering the elevated zone diameter breakpoints of the 2010 CLSI guidelines, 2/21 AmpC-positive strains were categorized as susceptible to extended-spectrum cephalosporins.
    Journal of clinical microbiology 06/2011; 49(8):2924-32. · 4.16 Impact Factor
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    ABSTRACT: In this prospective study all Enterobacteriaceae isolates (n = 2,129) recovered in the clinical microbiology laboratory during October 2009 to April 2010 were analyzed for AmpC production. Clinical and Laboratory Standards Institute (CLSI) cefoxitin and cefotetan susceptibility breakpoints and CLSI critical ESBL diameters were used to screen for potential AmpC producers. In total, 305 isolates (211 potential AmpC producers and 94 AmpC screen-negative isolates as a control group) were further analyzed by multiplex PCR for the detection of plasmid-encoded ampC beta-lactamase genes and by ampC promoter sequence analysis (considered as the gold standard). Cefoxitin and cefotetan were assessed as primary screening markers. The sensitivities of cefoxitin and cefotetan for the detection of AmpC production were 97.4 and 52.6%, respectively, and the specificities were 78.7 and 99.3%, respectively. As a phenotypic confirmation test, the Etest AmpC and the cefoxitin-cloxacillin double-disk synergy method (CC-DDS) were compared. The sensitivities for the Etest AmpC and the CC-DDS method were 77.4 and 97.2%, respectively, and the specificity was 100% for both methods. The results of the Etest AmpC were inconclusive for 10 isolates. With the CC-DDS method 2 inconclusive results were observed. Based on this study, we propose a comprehensive diagnostic flow chart for the detection of AmpC production consisting of a simple phenotypic screening and a single phenotypic confirmation test with inconclusive results being resolved by molecular analysis. For the proposed flow chart using (i) cefoxitin as a screening marker for AmpC production, (ii) the CC-DDS method as phenotypic confirmation, and (iii) molecular methods in case of inconclusive results, the sensitivity and specificity for AmpC detection would have been 97.4 and 100%, respectively, with respect to the studied isolates. The phenotypic methods used in the AmpC algorithm are simple to perform and easy to implement in the diagnostic laboratory.
    Journal of clinical microbiology 06/2011; 49(8):2798-803. · 4.16 Impact Factor
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    ABSTRACT: Lipoproteins are well known virulence factors of bacterial pathogens in general and of Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis, in particular. Lipoprotein lipidation between Gram-positive and Gram-negative bacteria differs significantly as these are di- and triacylated, respectively. Little is known about the lipid anchor of mycobacterial lipoproteins. We reported recently that mycobacterial LppX, a lipoprotein involved in synthesis of cell wall components is triacylated, although mycobacteria are classified as GC-rich Gram-positive bacteria. We here exploited the model organism Mycobacterium smegmatis for the expression of Mtb LprF and characterized N-terminal modifications at the molecular level. LprF is a putative lipoprotein of Mtb involved in signaling of potassium-dependent osmotic stress. LprF is extensively modified in a mycobacterium-specific manner by a thioether-linked diacylglyceryl residue with one ester-bound tuberculostearic- and one C16:0 fatty acid and additionally by a third N-linked C16:0 fatty acid, and a hexose.
    Biochemical and Biophysical Research Communications 11/2009; 391(1):679-84. · 2.28 Impact Factor
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    ABSTRACT: The Pax8(-/-) mouse provides an ideal animal model to study the consequences of congenital hypothyroidism, because its only known defect is the absence of thyroid follicular cells. Pax8(-/-) mice are, therefore, completely athyroid in postnatal life and die around weaning unless they are substituted with thyroid hormones. As reported recently, Pax8(-/-) mice can also be rescued and survive to adulthood by the additional elimination of the entire thyroid hormone receptor alpha (TRalpha) gene, yielding Pax8(-/-)TRalpha(o/o) double-knockout animals. This observation has led to the hypothesis that unliganded TRalpha1 might be responsible for the lethal phenotype observed in Pax8(-/-) animals. In this study we report the generation of Pax8(-/-)TRalpha1(-/-) double-knockout mice that still express the non-T(3)-binding TR isoforms alpha2 and Deltaalpha2. These animals closely resemble the phenotype of Pax8(-/-) mice, including growth retardation and a completely distorted appearance of the pituitary with thyrotroph hyperplasia and hypertrophy, extremely high TSH mRNA levels, reduced GH mRNA expression, and the almost complete absence of lactotrophs. Like Pax8(-/-) mice, Pax8(-/-)TRalpha1(-/-) compound mutants die around weaning unless they are substituted with thyroid hormones. These findings do not support the previous interpretation that the short life span of Pax8(-/-) mice is due to the negative effects of the TRalpha1 aporeceptor, but, rather, suggest a more complex mechanism involving TRalpha2 and an unliganded TR isoform.
    Endocrinology 08/2005; 146(7):3179-84. · 4.72 Impact Factor
  • Silke. Polsfuss
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    ABSTRACT: Hannover, Med. Hochsch., Diss., 2005.