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Objectives: CARB-ES-19 is a comprehensive, multicenter, nationwide study integrating whole-genome sequencing (WGS) in the surveillance of carbapenemase-producing K. pneumoniae (CP-Kpn) and E. coli (CP-Eco) to determine their incidence, geographical distribution, phylogeny, and resistance mechanisms in Spain. Methods: In total, 71 hospitals, representing all 50 Spanish provinces, collected the first 10 isolates per hospital (February to May 2019); CPE isolates were first identified according to EUCAST (meropenem MIC > 0.12 mg/L with immunochromatography, colorimetric tests, carbapenem inactivation, or carbapenem hydrolysis with MALDI-TOF). Prevalence and incidence were calculated according to population denominators. Antibiotic susceptibility testing was performed using the microdilution method (EUCAST). All 403 isolates collected were sequenced for high-resolution single-nucleotide polymorphism (SNP) typing, core genome multilocus sequence typing (cgMLST), and resistome analysis. Results: In total, 377 (93.5%) CP-Kpn and 26 (6.5%) CP-Eco isolates were collected from 62 (87.3%) hospitals in 46 (92%) provinces. CP-Kpn was more prevalent in the blood (5.8%, 50/853) than in the urine (1.4%, 201/14,464). The cumulative incidence for both CP-Kpn and CP-Eco was 0.05 per 100 admitted patients. The main carbapenemase genes identified in CP-Kpn were blaOXA−48 (263/377), blaKPC−3 (62/377), blaVIM−1 (28/377), and blaNDM−1 (12/377). All isolates were susceptible to at least two antibiotics. Interregional dissemination of eight high-risk CP-Kpn clones was detected, mainly ST307/OXA-48 (16.4%), ST11/OXA-48 (16.4%), and ST512-ST258/KPC (13.8%). ST512/KPC and ST15/OXA-48 were the most frequent bacteremia-causative clones. The average number of acquired resistance genes was higher in CP-Kpn (7.9) than in CP-Eco (5.5). Conclusion: This study serves as a first step toward WGS integration in the surveillance of carbapenemase-producing Enterobacterales in Spain. We detected important epidemiological changes, including increased CP-Kpn and CP-Eco prevalence and incidence compared to previous studies, wide interregional dissemination, and increased dissemination of high-risk clones, such as ST307/OXA-48 and ST512/KPC-3. [ABSTRACT FROM AUTHOR]

Objectives: CARB-ES-19 is a comprehensive, multicenter, nationwide study integrating whole-genome sequencing (WGS) in the surveillance of carbapenemase-producing K. pneumoniae (CP-Kpn) and E. coli (CP-Eco) to determine their incidence, geographical distribution, phylogeny, and resistance mechanisms in Spain. Methods: In total, 71 hospitals, representing all 50 Spanish provinces, collected the first 10 isolates per hospital (February to May 2019); CPE isolates were first identified according to EUCAST (meropenem MIC > 0.12 mg/L with immunochromatography, colorimetric tests, carbapenem inactivation, or carbapenem hydrolysis with MALDI-TOF). Prevalence and incidence were calculated according to population denominators. Antibiotic susceptibility testing was performed using the microdilution method (EUCAST). All 403 isolates collected were sequenced for high-resolution single-nucleotide polymorphism (SNP) typing, core genome multilocus sequence typing (cgMLST), and resistome analysis. Results: In total, 377 (93.5%) CP-Kpn and 26 (6.5%) CP-Eco isolates were collected from 62 (87.3%) hospitals in 46 (92%) provinces. CP-Kpn was more prevalent in the blood (5.8%, 50/853) than in the urine (1.4%, 201/14,464). The cumulative incidence for both CP-Kpn and CP-Eco was 0.05 per 100 admitted patients. The main carbapenemase genes identified in CP-Kpn were bla OXA–48 (263/377), bla KPC–3 (62/377), bla VIM–1 (28/377), and bla NDM–1 (12/377). All isolates were susceptible to at least two antibiotics. Interregional dissemination of eight high-risk CP-Kpn clones was detected, mainly ST307/OXA-48 (16.4%), ST11/OXA-48 (16.4%), and ST512-ST258/KPC (13.8%). ST512/KPC and ST15/OXA-48 were the most frequent bacteremia-causative clones. The average number of acquired resistance genes was higher in CP-Kpn (7.9) than in CP-Eco (5.5). Conclusion: This study serves as a first step toward WGS integration in the surveillance of carbapenemase-producing Enterobacterales in Spain. We detected important epidemiological changes, including increased CP-Kpn and CP-Eco prevalence and incidence compared to previous studies, wide interregional dissemination, and increased dissemination of high-risk clones, such as ST307/OXA-48 and ST512/KPC-3. [ABSTRACT FROM AUTHOR]

Objectives To analyse the ongoing epidemiology of KPC-producing Enterobacteriaceae after a non-ST258 KPC-3-producing Klebsiella pneumoniae outbreak in a university hospital in Madrid, Spain. Methods Enterobacterial isolates (one per patient based on bacterial identification and typing patterns) with carbapenem MICs higher than the EUCAST epidemiological cut-off values, a positive modified Hodge test and carbapenem/boronic acid combination disc test results were studied (16 March 2010 to 31 January 2012) and compared with KPC-producing isolates previously described in our institution (September 2009 to February 2010). The bacterial population structure (PFGE and multilocus sequence typing), carbapenemase genes and KPC plasmids were studied. Patients' clinical records were reviewed. Results Twenty-four KPC-producing Enterobacteriaceae (20 K. pneumoniae, 2 Escherichia coli and 2 Enterobacter cloacae) from 23 patients (13 males, median age 72 years) were studied. Most KPC-producer strains were considered as colonizers. Six K. pneumoniae clones (ST11, ST20, ST384, ST454, ST659 and ST971), two E. coli clones (ST224 and ST357) and one E. cloacae clone were identified. blaKPC-3 was located on IncFIIK plasmids of ∼100 kb (E. coli ST357 and K. pneumoniae ST384, ST454, ST659 and ST971 clones) and on IncN plasmids of ∼40 kb (K. pneumoniae ST11 clone). Non-typeable plasmids of ∼20 kb containing blaKPC-2 were detected in scarcely represented clones (K. pneumoniae ST20, E. coli ST224 and E. cloacae). Conclusions During the 2 year period following the emergence of non-ST258 KPC-3-producing K. pneumoniae isolates in our institution, the blaKPC-3 and blaKPC-2 genes efficiently penetrated other Enterobacteriaceae lineages. Non-ST258 K. pneumoniae isolates were mostly responsible for the dissemination of KPC enzymes, producing a complex epidemiological picture. [ABSTRACT FROM PUBLISHER]

Carbapenemase-producing Enterobacterales (CPE) have become an important public health concern. In our hospital, VIM enzymes were first detected in 2005, Klebsiella pneumoniae carbapenemase (KPC) enzymes in 2009, and OXA-48 enzymes in 2012. We assess the population biology of the first OXA-48-producing Enterobacterales isolates recovered in our hospital (2012 to 2013) where infections by other carbapenemases had been endemic for several years. Over a 21-month period, 71 isolates (61 Klebsiella pneumoniae , 5 Escherichia coli , 2 Klebsiella aerogenes , and 1 each of Enterobacter cloacae ,  Klebsiella oxytoca , and Citrobacter amalonaticus ) recovered from clinical and surveillance specimens from 57 patients (22.8% nonhospitalized) were investigated for OXA-48-like-producing enzymes. Analyses for characterization and determination of the location of the bla OXA-48 gene, plasmid transferability, sequence, and clonal relatedness were performed. Most of the isolates also coproduced CTX-M-15 (57/71, 80.3%) and/or VIM-1 (7/71, 9.8%). K. pneumoniae was predominantly identified as sequence type 11 (ST11) (63.4%) and ST405 (9.8%) and E. coli as ST540, ST1406, ST3163, and ST4301. The bla OXA-48 gene was part of Tn 1999.2 located at the tir gene of plasmids (ca. ≥50 kb) of the IncL/M group, also carrying bla VIM-1 and bla CTX-M-15 genes. We selected one ST11 K. pneumoniae isolate for whole-genome sequencing in which we studied the plasmid containing the bla OXA-48 gene. This plasmid was compared with indexed plasmids existing in NCBI database by the use of BRIG and MAUVE. Our data suggest a rapid spread of bla OXA-48 genes between commonly isolated high-risk clones of Enterobacterales species, frequently associated with antibiotic resistance. Moreover, the emergence of the multiresistant ST11 K. pneumoniae clone among nonhospitalized patients emphasizes the difficulty of preventing its dissemination into the community. IMPORTANCE We present results of microbiological analysis of the first Enterobacterales isolates that were isolated in 2012 in our institution expressing OXA-48 carbapenemase. This enzyme confers resistance to carbapenems, an important group of antibiotics widely used in the hospitals. OXA-48 carbapenemase is currently present in many parts of the world, but it is found particularly frequently in the Mediterranean area. It was disseminated at the Ramón y Cajal Hospital and found to be associated with a particular Klebsiella pneumoniae strain, so-called high-risk clone ST11, which was previously found in our institution in association with other enzymes such as CTX-M-15, VIM-1, and KPC-3. This clone might have acquired a plasmid harboring the bla OXA-48 gene. Our results point out the importance of local epidemiology in the dissemination and maintenance of multidrug-resistant bacteria., (Copyright © 2020 Gijón et al.)

Objectives: To evaluate the performance of the eazyplex(®) SuperBug CRE system, a loop-mediated isothermal amplification (LAMP)-based system, for confirming the presence of carbapenemases in addition to CTX-M-type ESBLs in previously genotypically and/or phenotypically characterized clinical Enterobacteriaceae isolates recovered in two centres in Spain., Methods: A collection of 94 carbapenemase-producing strains previously characterized by conventional PCR and sequencing and a total of 45 prospectively collected isolates with phenotypes compatible with the presence of a carbapenemase were tested with the eazyplex(®) SuperBug CRE system. In both cases, the presence of an ESBL was also assessed. Results were evaluated to establish the accuracy of this rapid LAMP-based system as well as to determine the concordance between all approaches., Results: The eazyplex(®) SuperBug CRE system correctly detected bla carbapenemase genes with or without blaCTX-M genes in 100% of the molecularly characterized strains. Absolute concordance (100%) was also observed in the case of isolates with phenotypes compatible with the presence of a carbapenemase with or without an ESBL inferred by susceptibility patterns and phenotypic inhibitory profiles. Determinations performed with the eazyplex(®) SuperBug CRE system took 15 min., Conclusions: The eazyplex(®) SuperBug CRE system proved to be a powerful tool for the detection of different carbapenemases as well as CTX-M-type ESBLs in Enterobacteriaceae with a rapid resolution time. The test has the high-performance parameters attributable to the sensitivity and specificity already demonstrated by LAMP-based assays. These results assure the usefulness of this test for routine rapid confirmation of carbapenemase-producing Enterobacteriaceae., (© The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)