Extended-spectrum cephalosporin-resistant Escherichia coli in community, specialized outpatient clinic and hospital settings in Switzerland

Objectives Resistance to extended-spectrum cephalosporins (ESCs) in Escherichia coli can be due to the production of ESBLs, plasmid-mediated AmpCs (pAmpCs) or chromosomal AmpCs (cAmpCs). Information regarding type and prevalence of β-lactamases, clonal relations and plasmids associated with the bla genes for ESC-R E. coli (ESC-R-Ec) detected in Switzerland is lacking. Moreover, data focusing on patients referred to the specialized outpatient clinics (SOCs) are needed. Methods We analysed 611 unique E. coli isolated during September-December 2011. ESC-R-Ec were studied with microarrays, PCR/DNA sequencing for blaESBLs, blapAmpCs, promoter region of blacAmpC, IS elements, plasmid incompatibility group, and also implementing transformation, aIEF, rep-PCR and MLST. Results The highest resistance rates were observed in the SOCs, whereas those in the hospital and community were lower (e.g. quinolone resistance of 22.6%, 17.2% and 9.0%, respectively; P = 0.003 for SOCs versus community). The prevalence of ESC-R-Ec in the three settings was 5.3% (n = 11), 7.8% (n = 22) and 5.7% (n = 7), respectively. Thirty isolates produced CTX-M ESBLs (14 were CTX-M-15), 5 produced CMY-2 pAmpC and 5 hyper-expressed cAmpCs due to promoter mutations. Fourteen isolates were of sequence type 131 (ST131; 10 with CTX-M-15). blaCTX-M and blaCMY-2 were associated with an intact or truncated ISEcp1 and were mainly carried by IncF, IncFII and IncI1plasmids. Conclusions ST131 producing CTX-M-15 is the predominant clone. The prevalence of ESC-R-Ec (overall 6.5%) is low, but an unusual relatively high frequency of AmpC producers (25%) was noted. The presence of ESC-R-Ec in the SOCs and their potential ability to be exchanged between hospital and community should be taken into serious consideratio

Outbreak investigation including molecular characterization of community associated methicillin-resistant Staphylococcus aureus in a primary and secondary school in Eastern Switzerland

At our tertiary children's hospital, infections with newly detected methicillin-resistant Staphylococcus aureus (MRSA) among children attending primary (age 6-12 years) and secondary school (age 13-16 years) nearly doubled in 2018 compared to previous years. This observation initiated an epidemiological outbreak investigation including phenotypic (susceptibility testing) and genotypic (whole genome sequencing) characterization of the isolates. In addition, a cross-sectional study was conducted to determine source of the outbreak, colonization frequency and to identify risk factors for transmission using a questionnaire. As a result, 49 individuals were detected with 57 corresponding isolates. Based on the case definition combined with whole genome sequencing, a core cluster was identified that shared common genetic features and a similar antimicrobial susceptibility pattern (efflux-mediated macrolide resistance, tetracycline susceptibility along with presence of Panton-Valentine leukocidin). Epidemiologic evaluation identified a distinct school as a common risk factor. However, the source of the clustered infections within that school could not be further specified. No further cases could be detected after decolonization of infected and colonized children

Multifactorial resistance mechanisms associated with resistance to ceftazidime-avibactam in clinical Pseudomonas aeruginosa isolates from Switzerland

BackgroundIncreasing reports of multidrug resistance (MDR) in clinical Pseudomonas aeruginosa have led to a necessity for new antimicrobials. Ceftazidime-avibactam (CZA) is indicated for use against MDR P. aeruginosa across a broad range of infection types and particularly those that are carbapenem resistant. This study sought to determine the molecular mechanisms of CZA and imipenem (IPM)-resistance in clinical P. aeruginosa isolates obtained from Swiss hospitals.MethodsClinical P. aeruginosa isolates were obtained from inpatients in three hospitals in Switzerland. Susceptibility was determined by either antibiotic disc testing or broth microdilution according to EUCAST methodology. AmpC activity was determined using cloxacillin and efflux activity was determined using phenylalanine-arginine β-naphthylamide, in agar plates. Whole Genome Sequencing was performed on 18 clinical isolates. Sequence types (STs) and resistance genes were ascertained using the Centre for Genomic Epidemiology platform. Genes of interest were extracted from sequenced isolates and compared to reference strain P. aeruginosa PAO1.ResultsSixteen different STs were identified amongst the 18 isolates in this study indicating a high degree of genomic diversity. No carbapenemases were detected but one isolate did harbor the ESBL blaPER-1. Eight isolates were CZA-resistant with MICs ranging from 16-64 mg/L, and the remaining ten isolates had either low/wildtype MICs (n=6; 1-2 mg/L) or elevated, but still susceptible, MICs (n=4; 4-8 mg/L). Ten isolates were IPM-resistant, seven of which had mutations resulting in truncations of OprD, and the remaining nine IPM-susceptible isolates had intact oprD genes. Within CZA-R isolates, and those with reduced susceptibility, mutations resulting in ampC derepression, OprD loss, mexAB overexpression and ESBL (blaPER-1) carriage were observed in various combinations and one harbored a truncation of the PBP4 dacB gene. Within the six isolates with wildtype-resistance levels, five had no mutations that would affect any antimicrobial resistance (AMR) genes of interest when compared to PAO1.ConclusionThis preliminary study highlights that CZA-resistance in P. aeruginosa is multifactorial and could be caused by the interplay between different resistance mechanisms including ESBL carriage, increased efflux, loss of permeability and derepression of its intrinsic ampC

Major structural features of IncM and IncL plasmids.

<p>p202c, R69, R471 and pKPN-El-Nr.7 plasmids sequenced in this study are compared with pNDM-OM from <i>K</i>. <i>pneumoniae</i> 601 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0123063#pone.0123063.ref007" target="_blank">7</a>] and pEL60 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0123063#pone.0123063.ref026" target="_blank">26</a>]. In the in scale schematic representation, the open reading frames identified in the sequence are represented with arrows of different colors: the <i>tra</i> and <i>trb</i> transfer loci are in green; the <i>traX</i>, <i>traY</i>, <i>excA</i> entry exclusion genes of the IncM type are in purple, whereas those of the IncL type are in pale blue; resistance genes are in red; transposon-related genes [<i>tnpA</i>, <i>tnpR</i>, <i>tnpM</i>], insertion sequences, integrase and resolvase genes are in yellow; the replicase gene is in blue; partitioning genes, toxin-antitoxin and other stabilization systems are in brown; additional genes of unknown function are in white. The line above each plasmid represents the core genome used in comparative analysis.</p

<i>Eco</i>RI, <i>Eco</i>RV and <i>Bam</i>HI restriction profiles of purified IncL/M plasmid DNAs, analyzed on 0.8% agarose gel.

<p>DNA on the gel: DNA ladder XV (Roche Diagnostic, GmbH, Mannheim, Germany); <i>E</i>. <i>coli</i> pKPN-El-Nr.7 plasmid; <i>E</i>. <i>coli</i> R69 (IncM reference plasmid); plasmid purified from the R69 x pKPN-El-Nr.7 transconjugant.</p

Homology trees of the ExcA, TraX, TraY protein sequences.

<p>The deduced protein sequences of the proteins from each respective plasmid were downloaded from the GenBank or deduced from the DNA sequences of plasmids performed in this study. The percentage of amino acid identity among the compared protein sequences is shown on each branch of the trees. Plasmids positive for the <i>bla</i>_OXA-48 gene are highlighted by pale blue panels.</p

Phylogenetic tree and nucleotide sequence alignment of the major stem-and-loop <i>inc</i>RNA of IncL and IncM plasmid.

<p>The <i>inc</i>RNA sequences were downloaded from the GenBank or identified in plasmids sequenced in this study. The unrooted phylogenetic tree inferred the evolutionary relationships among the various <i>inc</i>RNAs based upon similarities and differences in their nucleotide sequences. The part of the phylogeny tree corresponding to plasmids positive for the <i>bla</i>_OXA-48 gene is highlighted by pale blue panels. In the panel showing the nucleotide alignment, the residues showing 100% identity have been shaded. The white parts of the alignment show the mismatches identified among the IncL and IncM <i>inc</i>RNA sequences.</p