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Third-generation cephalosporin resistance (3GC-R) in Escherichia coli is a rising problem in human and farmed-animal populations. We conducted wholegenome sequencing analysis of 138 representative 3GC-R isolates previously collected from dairy farms in southwest England and confirmed by PCR to carry acquired 3GC-R genes. This analysis identified blaCTX-M (131 isolates encoding CTX-M-1, -14, -15, -and 32 and the novel variant CTX-M-214), blaCMY-2 (6 isolates), and blaDHA-1 (1 isolate). A highly conserved plasmid was identified in 73 isolates, representing 27 E. coli sequence types. This novel ~220-kb IncHI2 plasmid carrying blaCTX-M-32 was sequenced to closure and designated pMOO-32. It was found experimentally to be stable in cattle and human transconjugant E. coli even in the absence of selective pressure and was found by multiplex PCR to be present on 26 study farms representing a remarkable range of transmission over 1,500 square kilometers. However, the plasmid was not found among human urinary E. coli isolates we recently characterized from people living in the same geographical location, collected in parallel with farm sampling. There were close relatives of two blaCTX-M plasmids circulating among eight human and two cattle isolates, and a closely related blaCMY-2 plasmid was found in one cattle and one human isolate. However, phylogenetic evidence of recent sharing of 3GC-R strains between farms and humans in the same region was not found. [ABSTRACT FROM AUTHOR]

Objectives: Third-generation cephalosporin-resistant Escherichia coli from community-acquired urinary tract infections are increasingly reported worldwide. We sought to determine and characterize the mechanisms of cefotaxime resistance employed by urinary E. coli obtained from primary care, over 12 months, in Bristol and surrounding counties in South-West England.Methods: Cefalexin-resistant E. coli isolates were identified from GP-referred urine samples using disc susceptibility testing. Cefotaxime resistance was determined by subsequent plating onto MIC breakpoint plates. β-Lactamase genes were detected by PCR. WGS was performed on 225 isolates and analyses were performed using the Center for Genomic Epidemiology platform. Patient information provided by the referring general practices was reviewed.Results: Cefalexin-resistant E. coli (n=900) isolates were obtained from urines from 146 general practices. Following deduplication by patient approximately 69% (576/836) of isolates were cefotaxime resistant. WGS of 225 isolates identified that the most common cefotaxime-resistance mechanism was blaCTX-M carriage (185/225), followed by plasmid-mediated AmpCs (pAmpCs) (17/225), AmpC hyperproduction (13/225), ESBL blaSHV variants (6/225) or a combination of both blaCTX-M and pAmpC (4/225). Forty-four STs were identified, with ST131 representing 101/225 isolates, within which clade C2 was dominant (54/101). Ciprofloxacin resistance was observed in 128/225 (56.9%) of sequenced isolates, predominantly associated with fluoroquinolone-resistant clones ST131 and ST1193.Conclusions: Most cefalexin-resistant E. coli isolates were cefotaxime resistant, predominantly caused by blaCTX-M carriage. The correlation between cefotaxime resistance and ciprofloxacin resistance was largely attributable to the high-risk pandemic clones ST131 and ST1193. Localized epidemiological data provide greater resolution than regional data and can be valuable for informing treatment choices in the primary care setting. [ABSTRACT FROM AUTHOR]

Objectives: Carbapenemase-producing Enterobacteriaceae (CPE) have been increasingly reported in the UK since 2003. We analysed patient and isolate data for CPE confirmed by the national reference laboratory from laboratories in the West Midlands region from November 2007 to December 2014.Methods: MICs were determined by BSAC agar dilution methodology and isolates exhibiting resistance to one or more carbapenems were screened for carbapenemase genes by PCR. Plasmid analyses were performed after electro-transformation of carbapenemase-encoding plasmids. WGS was performed on both transformants and clinical isolates. Patient data provided by the sending laboratories were reviewed.Results: During the study period, CPE ( n  = 139) were submitted from 13 laboratories in the West Midlands region, originating from 108 patients and including one environmental isolate. CPE submissions increased significantly from 2009 onwards. Isolates were predominantly Klebsiella pneumoniae (89/139) obtained from inpatients. WGS was performed on all clinical isolates and transformants. After deduplication 119 isolates and 96 transformants remained for analysis. Within these, four families of carbapenemase genes were identified: bla NDM (69/119), bla KPC (26/119), bla OXA-48-like (16/119) and bla VIM (7/119); one isolate carried both bla NDM and bla OXA-48-like . Isolates represented diverse STs and plasmid replicon types. Plasmid analyses identified plasmids of different replicon types encoding bla KPC , bla NDM and bla OXA-48-like genes, found across several species and STs.Conclusions: CPE have been reported increasingly in the West Midlands region over a 7 year period. bla NDM , bla KPC and bla OXA-48-like were the dominant carbapenemase genes and were found in a range of diverse genomic/plasmid environments, highlighting their ability to mobilize across different plasmids, often impeding the detection of outbreaks. [ABSTRACT FROM AUTHOR]

Background: Early and accurate treatment of infections due to carbapenem-resistant organisms is facilitated by rapid diagnostics, but rare resistance mechanisms can compromise detection. One year after a Guiana Extended-Spectrum (GES)-5 carbapenemase-positive Klebsiella oxytoca infection was identified by whole-genome sequencing (WGS; later found to be part of a cluster of 3 cases), a cluster of 11 patients with GES-5-positive K. oxytoca was identified over 18 weeks in the same hospital., Methods: Bacteria were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry, antimicrobial susceptibility testing followed European Committee on Antimicrobial Susceptibility Testing guidelines. Ertapenem-resistant isolates were referred to Public Health England for characterization using polymerase chain reaction (PCR) detection of GES, pulsed-field gel electrophoresis (PFGE), and WGS for the second cluster., Results: The identification of the first GES-5 K. oxytoca isolate was delayed, being identified by WGS. Implementation of a GES-gene PCR informed the occurrence of the second cluster in real time. In contrast to PFGE, WGS phylogenetic analysis refuted an epidemiological link between the 2 clusters; it also suggested a cascade of patient-to-patient transmission in the later cluster. A novel GES-5-encoding plasmid was present in K. oxytoca, Escherichia coli, and Enterobacter cloacae isolates from unlinked patients within the same hospital group and in human and wastewater isolates from 3 hospitals elsewhere in the United Kingdom., Conclusions: Genomic sequencing revolutionized the epidemiological understanding of the clusters; it also underlined the risk of covert plasmid propagation in healthcare settings and revealed the national distribution of the resistance-encoding plasmid. Sequencing results also informed and led to the ongoing use of enhanced diagnostic tests for detecting carbapenemases locally and nationally., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.)