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We describe a modification of the most probable number (MPN) method for rapid enumeration of antimicrobial-resistant Escherichia coli bacteria in aqueous environmental samples. E. coli (total and antimicrobialresistant) bacteria were enumerated in effluent samples from a hospital (n ‫؍‬ 17) and municipal sewers upstream (n ‫؍‬ 5) and downstream (n ‫؍‬ 5) from the hospital, effluent samples from throughout the treatment process (n ‫؍‬ 4), and treated effluent samples (n ‫؍‬ 13). Effluent downstream from the hospital contained a higher proportion of antimicrobial-resistant E. coli than that upstream from the hospital. Wastewater treatment reduced the numbers of E. coli bacteria (total and antimicrobial resistant); however, antimicrobialresistant E. coli was not eliminated, and E. coli resistant to cefotaxime (including extended-spectrum betalactamase [ESBL] producers), ciprofloxacin, and cefoxitin was present in treated effluent samples.

Salmonellosis is the second most common cause of food-borne illness worldwide. Contamination of surfaces in food processing environments may result in biofilm formation with a risk of food contamination. Effective decontamination of biofilm-contaminated surfaces is challenging. Using the CDC biofilm reactor, the activities of sodium hypochlorite, sodium hydroxide, and benzalkonium chloride were examined against an early (48-h) and relatively mature (168-h) Salmonella biofilm. All 3 agents result in reduction in viable counts of Salmonella; however, only sodium hydroxide resulted in eradication of the early biofilm. None of the agents achieved eradication of mature biofilm, even at the 90-min contact time. Studies of activity of chemical disinfection against biofilm should include assessment of activity against mature biofilm. The difficulty of eradication of established Salmonella biofilm serves to emphasize the priority of preventing access of Salmonella to postcook areas of food production facilities.

BackgroundThis study examined colonisation with and characteristics of antimicrobial-resistant organisms among residents of a long-term care facility (LTCF) over one year, including strain persistence and molecular diversity among isolates of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae.MethodsSixty-four residents of a LTCF were recruited (51 at baseline, 13 during the year). Data on dependency levels, hospitalisations, and antimicrobial prescribing were collected. Nasal and rectal swabs and catheter urine specimens were examined quarterly, using chromogenic agars, for ESBL-producing Enterobacteriaceae, carbapenemase-producing Enterobacteriaceae (CPE), vancomycin-resistant enterococci (VRE), and meticillin-resistant S. aureus (MRSA). All ESBL-producing E. coli (ESBL-EC) were characterised by pulsed-field gel electrophoresis (PFGE) and PCR to assess for sequence type (ST) ST131, its resistance-associated H30 and H30-Rx subclones, and blaCTX-M,blaTEM,blaSHV, and blaOXA-1.ResultsThe overall number of residents colonised, by organism, was as follows: ESBL-EC, 35 (55%); MRSA, 17 (27%); ESBL-producing K. pneumoniae (ESBL-KP), 5 (8%); VRE, 2 (3%) and CPE, 0 (0%). All 98 ESBL-EC isolates were H30-Rx ST131, with blaCTX-M-group 1. By PFGE, a group of 91 ESBL-EC (from 33 participants) had ≥ 85% similar profiles and resembled UK epidemic strain A/ international pulsotype PFGE812. Sequential ESBL-EC from individual residents were closely related. Six ESBL-KP isolates, from five participants, had blaCTX-M-group 1 and by PFGE were closely related. Colonisation with ESBL and MRSA was associated with location within the LTCF and previous exposure to antimicrobials.ConclusionsAmong LTCF residents, colonisation with ESBL-EC and MRSA was common. All ESBL-EC were H30-Rx ST131, consistent with clonal dissemination.

Escherichia coli sequence type 131 (ST131) is a pandemic clone that is evolving rapidly with increasing levels of antimicrobial resistance. Here, we investigated an outbreak of E. coli ST131 producing extended spectrum β-lactamases (ESBLs) in a long-term care facility (LTCF) in Ireland by combining data from this LTCF (n=69) with other Irish (n=35) and global (n=690) ST131 genomes to reconstruct the evolutionary history and understand changes in population structure and genome architecture over time. This required a combination of short- and long-read genome sequencing, de novo assembly, read mapping, ESBL gene screening, plasmid alignment and temporal phylogenetics. We found that Clade C was the most prevalent (686 out of 794 isolates, 86 %) of the three major ST131 clades circulating worldwide (A with fimH41, B with fimH22, C with fimH30), and was associated with the presence of different ESBL alleles, diverse plasmids and transposable elements. Clade C was estimated to have emerged in c. 1985 and subsequently acquired different ESBL gene variants (bla CTX-M-14 vs bla CTX-M-15 ). An ISEcp1-mediated transposition of the bla CTX-M-15 gene further increased the diversity within Clade C. We discovered a local clonal expansion of a rare C2 lineage (C2_8) with a chromosomal insertion of bla CTX-M-15 at the mppA gene. This was acquired from an IncFIA plasmid. The C2_8 lineage clonally expanded in the Irish LTCF from 2006, displacing the existing C1 strain (C1_10), highlighting the potential for novel ESBL-producing ST131 with a distinct genetic profile to cause outbreaks strongly associated with specific healthcare environments.

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Antimicrobial resistance is a major public health concern. Carbapenemase-producing Enterobacterales (CPE) represent a significant health threat as some strains are resistant to almost all available antibiotics. The aim of this research was to examine hospital effluent and municipal wastewater in an urban area in Ireland for CPE. Samples of hospital effluent ( n = 5), municipal wastewater before (n = 5) and after ( n = 4) the hospital effluent stream joined the municipal wastewater stream were collected over a nine-week period (May–June 2017). All samples were examined for CPE by direct plating onto Brilliance CRE agar. Isolates were selected for susceptibility testing to 15 antimicrobial agents in accordance with EUCAST criteria. Where relevant, isolates were tested for carbapenemase-encoding genes by real-time PCR. CPE were detected in five samples of hospital effluent, one sample of pre-hospital wastewater and three samples of post-hospital wastewater. Our findings suggest hospital effluent is a major contributor to CPE in municipal wastewater. Monitoring of hospital effluent for CPE could have important applications in detection and risk management of unrecognised dissemination of CPE in both the healthcare setting and the environment.

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