Complete Nucleotide Sequence of a Citrobacter freundii Plasmid Carrying KPC-2 in a Unique Genetic Environment

The complete and annotated nucleotide sequence of a 54,036-bp plasmid harboring a blaKPC-2 gene that is clonally present in Citrobacter isolates from different species is presented. The plasmid belongs to incompatibility group N (IncN) and harbors the class A carbapenemase KPC-2 in a unique genetic environment

Dissemination of carbapenem-resistant Pseudomonas aeruginosa isolates and their susceptibilities to ceftolozane-tazobactam in Germany

Pseudomonas aeruginosa (PA) is a major cause of healthcare-associated infections. Antipseudomonal carbapenems are among the antimicrobial agents used to treat PA infections, but several mechanisms of resistance, including the production of a carbapenemase (CP), may compromise their clinical efficacy. The objectives of this study were to determine: (i) the dissemination of carbapenem-resistant CP-negative and CP-positive PA isolates; and (ii) the in-vitro activity of ceftolozane-tazobactam (CTT) against carbapenemsusceptible and carbapenem-resistant isolates. Isolates were collected prospectively from January 2016 to April 2017 at 20 German medical laboratories. Each centre was asked to provide 50 consecutive isolates from hospitalized patients. Overall, 985 isolates were collected, of which 34% were obtained from intensive care patients. Seven hundred and thirty-eight (74.9%) isolates were susceptible to both imipenem and meropenem (Subgroup I), and 247 (25.1%) isolates were resistant to carbapenems (Subgroup II): 125 (12.7%) were imipenem-resistant but meropenem-susceptible, 12 (1.2%) were meropenem-resistant but imipenem-susceptible, and 110 (11.2%) were resistant to both carbapenems (Subgroup III). A CP was detected in 28 (2.8%) isolates (predominantly VIM-2). Nine hundred and fifty (96.4%) isolates were CTTsusceptible. Susceptibility to CTT was seen in 99.6% of Subgroup I isolates, 87% of Subgroup II isolates and 74.5% of Subgroup III isolates. Overall, 2.8% of PA produced a CP, while 22.2% were carbapenemresistant, CP-non-producing isolates. Based on these findings, CTT may be considered for treatment of PA infections, particularly those caused by multi-drug-resistant CP-non-producing isolates. (C) 2020 Elsevier B.V. and International Society of Chemotherapy. All rights reserved

Comparison of VITEK (R) 2, three different gradient strip tests and broth microdilution for detecting vanB-positive Enterococcus faecium isolates with low vancomycin MICs

Objectives In 2018, EUCAST issued a warning regarding unreliable results of gradient strip tests for confirming vancomycin resistance in enterococci. We compared the performance of various diagnostic standard and confirmatory tests to identify and determine vanB-type vancomycin resistance. Methods We analysed a collection of vanB-positive Enterococcus faecium isolates (n=68) with low vancomycin MICs and compared the performance of VITEK (R) 2 (bioMerieux), broth microdilution and three gradient strip tests from different providers (Oxoid, Liofilchem and bioMerieux). For the latter we compared the standard procedure with a protocol with increased inoculum, a rich agar medium and a longer incubation time ('macromethod'). Results The sensitivity of VITEK (R) 2 was 81% compared with 72% for broth microdilution and 61%-63% for the three gradient strip tests using standard conditions. The macromethod substantially improved the performance of all strip tests resulting in a sensitivity of 89%-96% after 48h of incubation. Conclusions We recommend that EUCAST changes the present warning against the general use of MIC strips. When MIC strips are used to either exclude or confirm suspected vancomycin resistance in E. faecium, and a PCR is not available, the macromethod should be employed. For clinically relevant enterococci, where a rapid therapeutic decision is needed, a molecular test (e.g. PCR) should be favoured in order to save time and to further increase sensitivity

In vitro activity of cefiderocol against aerobic Gram-negative bacterial pathogens from Germany

Objectives: Cefiderocol (CID), also known as S-649266, a novel siderophore cephalosporin, possesses potent activity against multidrug-resistant aerobic Gram-negative bacteria (GNB). This study aimed to determine the in vitro activity of CID against two different sets of GNB: i) a random sample of 213 clinical isolates, including 17 extended-spectrum beta-lactamase (ESBL) producers, obtained from intensive care unit patients with nosocomial infections collected during a multicentre surveillance study (set I); and ii) a group of 59 challenge GNB producing various types of carbapenemases (CP; set II). Methods: Minimum inhibitory concentrations (MICs) were determined using the microdilution method according to the standard ISO 20776-1. Iron-depleted medium was used for testing CID. Results: CID inhibited 97.2% of set I isolates at the EUCAST susceptibility breakpoint of <= 2 mg/L. The concentrations of CID inhibiting 50% and 90% (MIC50/90) of the Enterobacterales isolates (n = 146) were 0.12/1.0 mg/L, with ESBL-positive isolates tending to exhibit higher MICs than ESBL-negative isolates to CID. MIC50/90 values of CID for isolates of the Acinetobacter baumannii group (n = 13) and Pseudomonas aeruginosa (n = 54) were 0.06/0.12 mg/L and 0.12/0.5 mg/L, respectively. Further, CID inhibited 88.1% of set II CP-producing isolates at <= 2 mg/L. All seven class D CP-producing Acinetobacter baumannii were inhibited at < 0.25 mg/L. MIC50/90 values for CP-producing Enterobacterales (n = 30) and Pseudomonas aeruginosa (n = 22) were 1/4 mg/L and 0.5/2 mg/L, respectively. Conclusion: CID showed potent activity against Acinetobacter baumannii, Enterobacterales and Pseudomonas aeruginosa, including CP-producing isolates. Overall, CID inhibited 259 of 272 (95.2%) GNB at <= 2 mg/L. (C) 2020 The Authors. Published by Elsevier Ltd

Comparison of Phenotypic Tests and an Immunochromatographic Assay and Development of a New Algorithm for Detection of OXA-48-like Carbapenemases

OXA-48 is the most prevalent carbapenemase in Enterobacteriaceae in Europe and the Middle East, but it is frequently missed because many isolates display low MICs for carbapenems. Furthermore, in contrast to metallo-beta-lactamases or Klebsiella pneumoniae carbapenemases (KPC), no specific inhibitor is available for the phenotypic detection of OXA-48. Molecular detection of blaOXA-48 is the gold standard but is not available in many laboratories. A few phenotypic assays have been described but have not been independently evaluated. The aim of this study was the systematic comparison of phenotypic tests and an immunochromatographic assay (ICT) for the detection of OXA-48/OXA-48-like carbapenemases and the development of an algorithm for reliable phenotypic detection of OXA-48. Four phenotypic tests (temocillin disk test, faropenem disk test, OXA-48 disk test, and high-inoculum [HI] OXA-48 disk test) and a new ICT (OXA-48 K-SeT) were compared by using a set of 166 Enterobacteriaceae isolates, including isolates producing OXA-48/OXA-48-like carbapenemases (n = 84) or Ambler class A and B carbapenemases (n = 41) and carbapenemase-negative isolates (n = 41). The sensitivity and specificity for the different assays were 100% and 43.9% for temocillin, 57.1% and 98.8% for faropenem, 53.6% and 100% for the OXA-48 disk test, 98.8% and 97.6% for the HI OXA-48 disk test, and 100% and 100% for the ICT, respectively. The ICT displayed the highest sensitivity and specificity and was the most rapid assay, but it is more costly than phenotypic assays. Based on these results, a new algorithm incorporating temocillin, faropenem, and ICT which allows cost-effective detection of OXA-48 with 100% sensitivity and specificity was developed

Population Dynamics among Methicillin-Resistant Staphylococcus aureus Isolates in Germany during a 6-Year Period

Methicillin-resistant Staphylococcus aureus (MRSA) originated from the health care setting but is now emerging in communities without health care contact (CA-MRSA) or in livestock (LA-MRSA). The impact on the whole MRSA population was assessed in a German prospective multicenter study. Thirty-three laboratories consecutively collected up to 50 MRSA isolates from infection or carriage during two sampling periods in 2004 to 2005 and 2010 to 2011. Patient-related data were collected using a standardized questionnaire. Methicillin resistance was confirmed by the detection of mecA or its homologue mecA(LGA251). The spa type and major virulence factors were analyzed for each isolate. In total, 1,604 (2004 to 2005) and 1,603 (2010 to 2011) MRSA isolates were analyzed; one isolate from each sampling period harbored mecA(LGA251). LA-MRSA increased significantly (odds ratio [OR] = 22.67, 95% confidence interval [CI] = 8.51 to 85.49, P < 0.0005) and spread over Germany, originating from northwestern regions. Panton-Valentine leukocidin-positive CA-MRSA rose significantly, particularly in southern Germany, but the proportion in 2010 to 2011 remained low (2.7%, OR = 2.80, 95% CI = 1.54 to 5.34, P < 0.0005). The emerging MRSA clones changed the MRSA population in Germany during a 6-year period significantly. The ongoing epidemiological shift and changes of MRSA sources create a need for revision of guidelines for MRSA infection control and treatment

Occurrence of carbapenemase-producing Klebsiella pneumoniae and Escherichia coli in the European survey of carbapenemase-producing Enterobacteriaceae (EuSCAPE): a prospective, multinational study

Background Gaps in the diagnostic capacity and heterogeneity of national surveillance and reporting standards in Europe make it difficult to contain carbapenemase-producing Enterobacteriaceae. We report the development of a consistent sampling framework and the results of the first structured survey on the occurrence of carbapenemase-producing Klebsiella pneumoniae and Escherichia coli in European hospitals. Methods National expert laboratories recruited hospitals with diagnostic capacities, who collected the first ten carbapenem non-susceptible clinical isolates of K pneumoniae or E coli and ten susceptible same-species comparator isolates and pertinent patient and hospital information. Isolates and data were relayed back to national expert laboratories, which made laboratory-substantiated information available for central analysis. Findings Between Nov 1, 2013, and April 30, 2014, 455 sentinel hospitals in 36 countries submitted 2703 clinical isolates (2301 [85%] Kpneurnoniae and 402 (15%) Ecoli). 850 (37%) of 2301 Kpneumoniae samples and 77 (19%) of 402 Ecoli samples were carbapenemase (KPC, NDM, OXA-48-like, or VIM) producers. The ratio of K pneumoniae to E coli was 11:1.1.3 patients per 10000 hospital admissions had positive clinical specimens. Prevalence differed greatly, with the highest rates in Mediterranean and Balkan countries. Carbapenemase-producing K pneumoniae isolates showed high resistance to last-line antibiotics. Interpretation This initiative shows an encouraging commitment by all participants, and suggests that challenges in the establishment of a continent-wide enhanced sentinel surveillance for carbapenemase-producing Entero-bacteriaeceae can be overcome. Strengthening infection control efforts in hospitals is crucial for controlling spread through local and national health care networks

Epidemic of carbapenem-resistant Klebsiella pneumoniae in Europe is driven by nosocomial spread

Public health interventions to control the current epidemic of carbapenem-resistant Klebsiella pneumoniae rely on a comprehensive understanding of its emergence and spread over a wide range of geographical scales. We analysed the genome sequences and epidemiological data of &gt;1,700 K. pneumoniae samples isolated from patients in 244 hospitals in 32 countries during the European Survey of Carbapenemase-Producing Enterobacteriaceae. We demonstrate that carbapenemase acquisition is the main cause of carbapenem resistance and that it occurred across diverse phylogenetic backgrounds. However, 477 of 682 (69.9%) carbapenemase-positive isolates are concentrated in four clonal lineages, sequence types 11,15,101, 258/512 and their derivatives. Combined analysis of the genetic and geographic distances between isolates with different beta-lactam resistance determinants suggests that the propensity of K. pneumoniae to spread in hospital environments correlates with the degree of resistance and that carbapenemase-positive isolates have the highest transmissibility. Indeed, we found that over half of the hospitals that contributed carbapenemase-positive isolates probably experienced within-hospital transmission, and interhospital spread is far more frequent within, rather than between, countries. Finally, we propose a value of 21 for the number of single nucleotide polymorphisms that optimizes the discrimination of hospital clusters and detail the international spread of the successful epidemic lineage, ST258/512

Occurrence of carbapenemase-producing Klebsiella pneumoniae and Escherichia coli in the European survey of carbapenemase-producing Enterobacteriaceae (EuSCAPE): a prospective, multinational study

Background Gaps in the diagnostic capacity and heterogeneity of national surveillance and reporting standards in Europe make it difficult to contain carbapenemase-producing Enterobacteriaceae. We report the development of a consistent sampling framework and the results of the first structured survey on the occurrence of carbapenemase-producing Klebsiella pneumoniae and Escherichia coli in European hospitals. Methods National expert laboratories recruited hospitals with diagnostic capacities, who collected the first ten carbapenem non-susceptible clinical isolates of K pneumoniae or E coli and ten susceptible same-species comparator isolates and pertinent patient and hospital information. Isolates and data were relayed back to national expert laboratories, which made laboratory-substantiated information available for central analysis. Findings Between Nov 1, 2013, and April 30, 2014, 455 sentinel hospitals in 36 countries submitted 2703 clinical isolates (2301 [85%] Kpneurnoniae and 402 (15%) Ecoli). 850 (37%) of 2301 Kpneumoniae samples and 77 (19%) of 402 Ecoli samples were carbapenemase (KPC, NDM, OXA-48-like, or VIM) producers. The ratio of K pneumoniae to E coli was 11:1.1.3 patients per 10000 hospital admissions had positive clinical specimens. Prevalence differed greatly, with the highest rates in Mediterranean and Balkan countries. Carbapenemase-producing K pneumoniae isolates showed high resistance to last-line antibiotics. Interpretation This initiative shows an encouraging commitment by all participants, and suggests that challenges in the establishment of a continent-wide enhanced sentinel surveillance for carbapenemase-producing Entero-bacteriaeceae can be overcome. Strengthening infection control efforts in hospitals is crucial for controlling spread through local and national health care networks