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Summary Background The global prevalence of extended-spectrum beta-lactamase-producing Escherichia coli is rising and is dominated by blaCTX-M spread by plasmids. Travellers to South Asia from Western Europe have high rates of acquisition of faecal CTX-M-producing E. coli (CTX-M-EC). Aims To determine the conjugative ability of CTX-M-EC acquired by healthy volunteers after travel to South Asia, the proportion of travel-acquired CTX-M-EC where blaCTX-M is encoded on a plasmid vs on the bacterial chromosome, and the relatedness of travel-acquired CTX-M-EC plasmids to previously sequenced plasmids. Methods Faecal samples were collected pre- and post-travel from 23 volunteers who visited South Asia, and CTX-M-EC were cultured. After short- and long-read sequencing, 10 plasmid sequences were identified and compared with previously sequenced plasmids in GenBank. Conjugation to E. coli K-12 was undertaken using filter mating. Findings Thirty-five percent of CTX-M-EC isolates tested transferred the blaCTX-M plasmid by conjugation. Travel-acquired CTX-M-EC carried blaCTX-M on a plasmid in 62% of isolates, whereas 38% of isolates had blaCTX-M on the chromosome. CTX-M-EC plasmids acquired after travel to South Asia had close homology to previously described epidemic plasmids which are widely disseminated in humans, animals and the natural environment. Conclusion Globally successful epidemic plasmids are involved in the spread of CTX-M-EC. Targeted strategies may be used to displace such plasmids from the host strain as part of efforts in infection prevention and control in healthcare settings. Bacteria with blaCTX-M plasmids were readily acquired by healthy volunteers, and were carried on return to the UK, providing opportunities for onward dissemination.

Summary Background Pseudomonas aeruginosa is a common opportunistic pathogen and molecular typing in outbreaks has linked patient acquisition to contaminated hospital water systems. Aim To elucidate the role of P. aeruginosa transmission rates in non-outbreak augmented care settings in the UK. Methods Over a 16-week period, all water outlets in augmented care units of four hospitals were sampled for P. aeruginosa and clinical isolates were collected. Outlet and clinical P. aeruginosa isolates underwent whole-genome sequencing (WGS), which with epidemiological data identified acquisition from water as definite (level 1), probable (level 2), possible (level 3), and no evidence (level 4). Findings Outlets were positive in each hospital on all three occasions: W (16%), X (2.5%), Y (0.9%) and Z (2%); and there were 51 persistently positive outlets in total. WGS identified likely transmission (at levels 1, 2 and 3) from outlets to patients in three hospitals for P. aeruginosa positive patients: W (63%), X (54.5%) and Z (26%). According to the criteria (intimate epidemiological link and no phylogenetic distance), approximately 5% of patients in the study ‘definitely’ acquired their P. aeruginosa from their water outlets in the intensive care unit. This study found extensive evidence of transmission from the outlet to the patients particularly in the newest hospital (W), which had the highest rate of positive outlets. Conclusions The overall findings suggest that water outlets are the most likely source of P. aeruginosa nosocomial infections in some settings, and that widespread introduction of control measures would have a substantial impact on infections.

Infections caused by antimicrobial-resistant bacterial pathogens are fast becoming an important global health issue. Strains of Escherichia coli are common causal agents of urinary tract infection and can carry multiple resistance genes. This includes the gene bla(CTX-M-15), which encodes an extended-spectrum beta-lactamase (ESBL). While studying antimicrobial resistance (AMR) in the environment, we isolated several strains of E. coli ST131 downstream of a wastewater treatment plan (WWTP) in a local river. These isolates were surviving in the river sediment, and characterization proved that a multiresistant phenotype was evident. Here, we show that E. coli strain 48 (river isolate ST131) provided a protective effect against a third-generation cephalosporin (cefotaxime) for susceptible E. coli strain 33 (river isolate ST3576) through secretion of a functional ESBL into the growth medium. Furthermore, extracellular ESBL activity was stable for at least 24 h after secretion. Proteomic and molecular genetic analyses identified CTX-M-15 as the major secreted ESBL responsible for the observed protective effect. In contrast to previous studies, outer membrane vesicles (OMVs) were not the route for CTX-M-15 secretion. Indeed, mutation of the type I secretion system led to a significant reduction in the growth of the ESBL-producing strain as well as a significantly reduced ability to confer protective effect. We speculate that CTX-M-15 secretion, mediated through active secretion using molecular machinery, provides a public goods service by facilitating the survival of otherwise susceptible bacteria in the presence of cefotaxime.

Infections caused by antimicrobial resistant bacterial pathogens are fast becoming an important global health issue. Strains of Escherichia coli are common causal agents of urinary tract infection and can carry multiple resistance genes. This includes the gene blaCTX-M-15 that encodes for an extended spectrum beta-lactamase (ESBL). While studying antimicrobial resistance (AMR) in the environment we isolated several strains of E. coli ST131 downstream of a WWTP in a local river. These isolates were surviving in the river sediment and characterisation proved that a multi-resistant phenotype was evident. Here, we show that E. coli strain 48 (river isolate ST131), provided a protective effect against a third-generation cephalosporin (cefotaxime) for a susceptible E. coli strain 33 (river isolate ST3576) through secretion of a functional ESBL into the growth medium. Furthermore, extracellular ESBL activity was stable for at least 24 h after secretion. Proteomic and molecular genetic analyses identified CTX-M-15 as the major secreted ESBL responsible for the observed protective effect. In contrast to previous studies, OMVs were not the sole route for CTX-M-15 secretion. Indeed, mutation of the Type I secretion system led to a significant reduction in the growth of the ESBL-producing strain as well as a significantly reduced ability to confer protective effect. We speculate that CTX-M-15 secretion, mediated through active secretion using molecular machinery provides a public goods service by facilitating the survival of otherwise susceptible bacteria in the presence of cefotaxime.Abstract importanceInfections caused by antimicrobial resistant bacterial pathogens have become an important global health issue. Wastewater treatment plants (WWTPs) have been identified as hotspots for the dissemination of antimicrobial resistant genes/bacteria into the environment. In this study, we investigated resistance enzyme secretion by a multi-drug resistant human pathogenic E. coli, isolated from a UK river, downstream of a WWTP. We present evidence that the resistant strain actively secreted an important resistance enzyme into the surrounding medium which degraded the antibiotic cefotaxime. This research provided evidence for the mechanism for secretion of this enzyme which could indicate a new target to tackle antibiotic resistance pathogens.

The Liverpool epidemic strain (LES) is an important transmissible clonal lineage ofPseudomonas aeruginosathat chronically infects the lungs of people with cystic fibrosis (CF). Previous studies have focused on the genomics of the LES in a limited number of isolates, mostly from one CF centre in the UK, and from studies highlighting identification of the LES in Canada. Here we significantly extend the current LES genome database by genome sequencing 91 isolates from multiple CF centres across the UK, and we describe the comparative genomics of this large collection of LES isolates from the UK and Canada. Phylogenetic analysis revealed that the 145 LES genomes analysed formed a distinct clonal lineage when compared with the widerP. aeruginosapopulation. Notably, the isolates formed two clades: one associated with isolates from Canada, and the other associated with UK isolates. Further analysis of the UK LES isolates revealed clustering by clinic geography. Where isolates clustered closely together, the association was often supported by clinical data linking isolates or patients. When compared with the earliest known isolate, LESB58 (from 1988), many UK LES isolates shared common loss-of-function mutations, such as in genesgltRandfleR. Other loss-of-function mutations identified in previous studies as common adaptations during CF chronic lung infections were also identified in multiple LES isolates. Analysis of the LES accessory genome (including genomic islands and prophages) revealed variations in the carriage of large genomic regions, with some evidence for shared genomic island/prophage complement according to clinic location. Our study reveals divergence and adaptation during the spread of the LES, within the UK and between continents.

Introduction Although faecal microbiota transplantation (FMT) appears to hold therapeutic potential for ulcerative colitis (UC), the optimal administration route and dose of FMT is unknown. This pilot trial aimed to identify the optimal route of administration to further test in an RCT. Methods In this prospective, three-centre, open-label, randomised study (STOP-Colitis pilot), we compared delivery of FMT via the naso-gastric (NG) or colonic (COLON) route in adult patients with active UC. Participants were administered 8 infusions of FMT over an 8 week period. Clinical response was defined as ≥3 point and ≥30% reduction in Mayo score at week 8 compared to baseline. Clinical remission was defined as Mayo score of ≤2, with no subscore >1 at week 8. The primary outcome was based on clinical response and safety at weeks 8 and 12, along with qualitative assessment of acceptability. Results 30 participants were randomised between March 2018 and April 2019; 16 to NG; 14 to COLON. 8 in NG arm and 2 patients in the COLON arm withdrew from the study before completion. Clinical response was achieved in more participants who received FMT via COLON compared with NG (9/12 [75%] vs 2/8 [25%]; adjusted relative risk [RR] 2.94 [95% CI, 0.84, 10.30]). Clinical remission was observed in more participants undergoing FMT via COLON compared to NG (6/12 [50%] vs 2/8 [25%] respectively; RR 1.89 [95% CI, 0.51, 6.99]). IBDQ and SF-36 scores at week 8 and 12 were similar in NG and COLON groups. Qualitative analysis showed greater patient and clinician acceptability for colonic delivery. There were three serious adverse events (one considered a serious adverse event) in 2 participants in the NG arm, and none in the COLON arm. Conclusion This pilot study suggests that in patients with active UC, FMT delivered via the COLON route appears to be safe and better tolerated with signals suggesting greater efficacy compared to the NG route. A randomised, double-blind, placebo-controlled trial of colonic delivery of FMT is now underway to determine clinical efficacy and safety.

Recent evidence suggests that anthropogenic activity can increase the levels of antimicrobial resistance (AMR) in the environment. Rivers and waterways are significant examples of environmental settings that have become repositories of antibiotics and antibiotic resistance genes (ARGs). Our recent study quantified drug concentrations in freshwater samples taken at a range of sites located on the Thames catchment; the highest levels of antibiotics and other drugs were recorded downstream of waste water treatment plants (WWTPs). One specific antibiotic: Trimethoprim (TMP) was shown at elevated concentrations reaching 2000ng/L at particular sites. We have also shown a correlative relationship between the residue of TMP and the prevalence of sulfonamide antibiotic resistance genes such as sul1. Despite this, there is still no evidence of a causative relationship between TMP concentrations and the prevalence of the ARGs at river sites. The aim of the current study was to conduct in-depth analysis using a combination of large metagenomic, geospatial and chemical datasets, in order to conduct a comparison between those sites with the highest TMP and lowest TMP levels across the Thames catchment. We aimed to establish the proximity of these sites to WWTPs, their population equivalence (PE) and land coverage. A secondary aim was to investigate seasonal variation in TMP and ARGs. Exploring these factors will help to decipher the clinical relevance of ARG accumulation at river sites. A significant correlation was shown between TMP levels at river sites and their distance downstream from a WWTP. Three sites located on the Rivers Cut and Ray showed significantly higher TMP concentrations in winter compared to summer. The population equivalence (PE) for sites with the highest TMP levels was significantly higher than those with the lowest levels. The land coverage of sites with the highest TMP levels was significantly more urban/suburban than sites with the lowest TMP concentrations, which were found to be significantly more arable. Five ARGs relevant to TMP and sulfonamides were identified across the Thames catchment. The most prevalent ARG was sul1, which was significantly more prevalent in winter compared to summer. By contrast sul2 was found to be significantly more prevalent in summer compared to winter at a site on the River Coln. The prevalence of the class 1 integron marker gene (inti1) did not differ significantly by season or between sites with the highest/lowest TMP levels.

Clostridioides difficile infection (CDI) is a common cause of nosocomial diarrhea and can sometimes lead to pseudo-membranous colitis and toxic megacolon. We previously reported that the PCR ribotype 002 was a common C. difficile ribotype in Hong Kong that was associated with increased mortality. In this study, we assessed in vitro bacteriological characteristics and in vivo virulence of ribotype 002 compared to other common ribotypes, including ribotypes 012, 014 and 046. We observed significantly higher toxin A (p

Introduction: Faecal microbiota transplantation (FMT) is an effective and licensed treatment for recurrent and refractory Clostridium difficile infection (CDI) and has shown encouraging signals for treatment of ulcerative colitis (UC). Access to FMT has been limited by the introduction of new regulations in the UK. Centres producing FMT are now required to hold a manufacturing licence from the Medicines and Healthcare products Regulatory Agency (MHRA). Methods:The first licenced FMT service in UK - University of Birmingham Microbiome Treatment Centre (UoBMTC) was launched in 2017. Policies and procedures were developed in accordance with MHRA ‘Good Manufacturing Practice’ guide. Results: Since August 2018, 132 FMT aliquots have been supplied for recurrent and refractory CDI to 39 NHS Trusts across UK. Twenty-nine of these Trusts did not have access / perform FMT prior to this service. In all cases, FMT was delivered within 48 hours (unless delayed delivery was requested). The service is the sole supplier of FMT since 2018 via an NHS Innovation and Technology Tariff. UoBMTC has also to date supplied 360 FMT aliquots for a multi-centre trial of FMT in UC (STOP-Colitis). Furthermore, the service has provided FMT to other refractory conditions within clinical settings. Conclusions: Development of a licenced FMT facility has greatly enhanced equality of access for this treatment across the NHS. FMT is provided at a zero-cost model for CDI and is available within 48 hours of request. UoBMTC continues to facilitate research in this field by providing FMT for clinical trials exploring its use for other indications.

The faecal, intestinal and mucosal microbiome, and manipulation of it, is the subject of intense categorisation, collation and study. Advances have been aided by novel culture-independent analytical techniques and a focus on how aberrations in the microbiome are associated with many diseases. Dramatic manipulation of the microbiome is possible using faecal microbiota transplantation (FMT). FMT has been shown to have a definite clinical role in treating recurrent Clostridium difficile infection, and there is accumulating clinical data for a possible role in treating ulcerative colitis. FMT preparation can only take place in a laboratory with licensed facilities and equipment allowing safe treatment and processing of appropriate donor samples. The future of FMT treatment may involve diseases of extra-intestinal origin. This review clarifies recent developments in research and provides a how-to guide for interested clinicians.

Anthropogenic inputs increase levels of antimicrobial resistance (AMR) in the environment, however, it is unknown how these inputs create this observed increase, and if anthropogenic sources impact AMR in environmental bacteria. The aim of this study was to characterise the role of waste water treatment plants (WWTPs) in the dissemination of class 1 integrons (CL1s) in the riverine environment. Using sample sites from upstream and downstream of a WWTP, we demonstrate through isolation and culture-independent analysis that WWTP effluent significantly increases both CL1 abundance and antibiotic resistance in the riverine environment. Characterisation of CL1-bearing isolates revealed that CL1s were distributed across a diverse range of bacteria, with identical complex genetic resistance determinants isolated from both human-associated and common environmental bacteria across connected sites. Over half of sequenced CL1s lacked the 3′-conserved sequence ('atypical’ CL1s); surprisingly, bacteria carrying atypical CL1s were on average resistant to more antibiotics than bacteria carrying 3′-CS CL1s. Quaternary ammonium compound (QAC) resistance genes were observed across 75% of sequenced CL1 gene cassette arrays. Chemical data analysis indicated high levels of boron (a detergent marker) downstream of the WWTP. Subsequent phenotypic screening of CL1-bearing isolates demonstrated that ~90% were resistant to QAC detergents, with in vitro experiments demonstrating that QACs could solely select for the transfer of clinical antibiotic resistance genes to a naive Escherichia coli recipient. In conclusion, this study highlights the significant impact of WWTPs on environmental AMR, and demonstrates the widespread carriage of clinically important resistance determinants by environmentally associated bacteria.

Objectives Although carbapenem susceptibility testing has been recommended for all Enterobacteriaceae from clinical specimens, for practical reasons a carbapenem is not included in many primary antibiotic panels for urine specimens. The 'iCREST' study sought carbapenemase-producing Enterobacteriaceae (CPE) in routine urine specimens yielding Gram-negative growth in five diagnostic laboratories in the UK. We sought also to compare locally and centrally determined MICs of meropenem and ceftazidime/avibactam. Methods Positive growth from up to 2000 urine specimens per laboratory was plated onto chromID® CARBA SMART agar. Suspected CPE colonies were tested locally by Etest for susceptibility to meropenem and ceftazidime/avibactam, and referred to central laboratories for PCR confirmation of CPE status and microbroth MIC determination. Results Twenty-two suspected CPE were identified from 7504 urine specimens. Ten were confirmed by PCR to have NDM (5), IMP (2), KPC (2) or OXA-48-like (1) carbapenemases. Locally determined ceftazidime/avibactam MICs showed complete categorical agreement with those determined centrally by microbroth methodology. The seven ceftazidime/avibactam-resistant isolates (MICs ≥256 mg/L) had NDM or IMP metallo-carbapenemases. Conclusions The frequency of confirmed CPE among Gram-negative urinary isolates was low, at 0.13% (10/7504), but CPE were found in urines at all five participating sites and the diversity of carbapenemase genes detected reflected the complex epidemiology of CPE in the UK. These data can inform local policies about the cost-effectiveness and clinical value of testing Gram-negative bacteria from urine specimens routinely against a carbapenem as part of patient management and/or infection prevention and control strategies.

Cross-sectional studies suggest an increasing trend in incidence and relatively low recurrence rates of Clostridium difficile infections in Asia than in Europe and North America. The temporal trend of C. difficile infection in Asia is not completely understood. We conducted a territory-wide population-based observational study to investigate the burden and clinical outcomes in Hong Kong, China, over a 9-year period. A total of 15,753 cases were identified, including 14,402 (91.4%) healthcare-associated cases and 817 (5.1%) community-associated cases. After adjustment for diagnostic test, we found that incidence increased from 15.41 cases/100,000 persons in 2006 to 36.31 cases/100,000 persons in 2014, an annual increase of 26%. This increase was associated with elderly patients, for whom incidence increased 3-fold over the period. Recurrence at 60 days increased from 5.7% in 2006 to 9.1% in 2014 (p

Limited data are available on temocillin susceptibilities in Enterobacteriaceae from Asian countries where antimicrobial resistance is prevalent. The in vitro activities of temocillin and 15 commonly used antimicrobials against 613 non-duplicate blood (n = 310) and urine (with clinically significant bacteriuria; n = 303) isolates of Enterobacteriaceae from patients who attended 3 out of 7 clusters of public hospitals of the Hospital Authority, Hong Kong, during 2015/2016 were tested. Minimum inhibitory concentrations (MICs) were determined by Clinical and Laboratory Standards Institute (CLSI) microbroth dilution (agar dilution with fosfomycin). For temocillin, MICs were also obtained using the British Society of Antimicrobial Chemotherapy (BSAC) microbroth dilution method and interpreted using the BSAC breakpoints. Overall, 93.0% (570) isolates were susceptible to temocillin using BSAC systemic breakpoint (≤8 mg/L) and all except 2 isolates were susceptible using the urinary breakpoint (≤32 mg/L). The extended spectrum beta-lactamase (ESBL) positivity rate was 23.2% (118 out of 508 E. coli, Klebsiella spp., Proteus spp.). Temocillin resistance rate to ESBL-positive isolates was 16.1% using the systemic breakpoint of ≤8 mg/L (MIC50 and MIC90 were 8 mg/L and 16 mg/L respectively). Two isolates (1 E. coli, temocillin MIC 64 mg/L, 1 Klebsiella sp., MIC 32 mg/mL) were resistant to meropenem and possessed the NDM-5 and KPC-2 genes respectively. Other susceptibility rates were: amoxicillin/clavulanate (59.1%), trimethoprim/sulfamethoxazole (62.5%), ciprofloxacin (71.5%), ceftriaxone (75.4%), nitrofurantoin (76.4%), gentamicin (78.3%), cefepime (81.1%), ceftazidime (83.5%), piperacillin/tazobactam (86%), colistin (88.8%), tigecycline (89.4%), fosfomycin (92.8%), ertapenem (99.0%), amikacin (99.2%) and meropenem (99.7%). Temocillin may be a useful alternative for the treatment of infections caused by ESBL and multi-drug-resistant Enterobacteriaceae in Hong Kong, particularly as resistance rates to ciprofloxacin, nitrofurantoin and piperacillin/tazobactam are high. Electronic supplementary material The online version of this article (10.1007/s10096-017-3091-8) contains supplementary material, which is available to authorized users

Globally, rates of ESBL-producing Enterobacteriaceae are rising. We undertook a literature review, and present the temporal trends in blaCTX-M epidemiology, showing that blaCTX-M-15 and blaCTX-M-14 have displaced other genotypes in many parts of the world. Explanations for these changes can be attributed to: (i) horizontal gene transfer (HGT) of plasmids; (ii) successful Escherichia coli clones; (iii) ESBLs in food animals; (iv) the natural environment; and (v) human migration and access to basic sanitation. We also provide explanations for the changing epidemiology of blaCTX-M-2 and blaCTX-M-27. Modifiable anthropogenic factors, such as poor access to basic sanitary facilities, encourage the spread of blaCTX-M and other antimicrobial resistance (AMR) genes, such as blaNDM, blaKPC and mcr-1. We provide further justification for novel preventative and interventional strategies to reduce transmission of these AMR genes.

Introduction Faecal Microbiota Transplant (FMT) is an efficacious treatment for recurrent Clostridium difficile infection (CDI) and has shown encouraging signals for inflammatory bowel disease. Access to FMT services in the NHS has been limited by the regulation of FMT as a medicinal product in the UK. To enable inter-institution supply, FMT manufacturers are required to hold a Specials licence from The Medicines and Healthcare products Regulatory Agency. Methods In 2017 the first licenced FMT service in UK was created at the University of Birmingham Microbiome Treatment Centre. Policies and procedures were developed using the Rules and Guidance for Pharmaceutical Manufacturers and Distributors (Orange Guide), which were used and maintained within a quality management system. The frozen FMT bank utilises anonymous unrelated healthy donors and provides FMT on a named patient basis for the treatment of CDI and supplies clinical trials via an investigational medicinal product (IMP) licence. Results Since August 2018, 53 faecal microbiota transplants have been supplied for recurrent and refractory CDI to 21 NHS Trusts across England and Wales. Twelve of these Trusts did not have access or perform FMT prior to the introduction of this service. In 98% (52/53) of cases FMT was delivered either within 48 hours or on the day of request if >48 hours from request. Clinical response to treatment was observed in 76% (16/21) of patients, with 88% (14/16) of these patients remaining symptom free at 90 days. In 2018 the service became the sole supplier of FMT under the 201–019 NHS Innovation and Technology Tariff. Under the IMP licence the service has successfully supplied over 360 faecal microbiota transplants for the pilot stage of a multi-centre trial of FMT in UC (STOP-Colitis). Conclusions Development of a licenced FMT facility at the University of Birmingham has improved equality of access to this novel treatment across the NHS. FMT has been provided within a zero cost model for CDI and is available generally within 48 hours of request. Patients with CDI benefit from improved care from this service, which limits morbidity and mortality from this severe infection. Supply via an IMP licence has enabled the required FMT infrastructure to be developed to support prospective FMT research in the UK.

Background: Myeloma causes profound immunodeficiency and recurrent, serious infections. There are approximately 5,500 new UK cases of myeloma per annum; a quarter will have a serious infection within 3 months of diagnosis. Newly diagnosed patients may benefit from antibiotic prophylaxis to prevent infection and early death but this may be associated with healthcare-associated infections (HCAI). Methods: The TEAMM trial was a multicentre randomised, double-blind, placebo-controlled trial in newly-diagnosed myeloma patients randomised to receive Levofloxacin or placebo for 12 weeks at the start of anti-myeloma treatment. Follow-up was 4-weekly to 16 weeks and again at 1 year. The primary outcome was time to first febrile episode or death in the first 12 weeks from start of trial treatment. Secondary outcomes included number of infections, deaths, healthcare-associated organism carriage and invasive infections, and overall survival (OS). Findings: 977 patients were randomised (489 levofloxacin, 488 placebo). Primary events (febrile episodes, deaths, febrile episodes with death) in levofloxacin versus placebo arms were 19% vs 27% (87, 4, 4 vs 112, 15, 7), respectively; HR=0.66 (95% CI 0.51-0.86) p=0.002. There was no difference for new acquisitions of clostridium difficile, Methicillin-resistant Staphylococcus aureus and Extended Spectrum Beta Lactamase-positive (ESBL) Gram-negative coliforms when assessed up to 16 weeks. There was an OS benefit at 12 weeks (p=0.008) but not at one year (p=0.94). Interpretation: The addition of prophylactic levofloxacin to active myeloma treatment during the first 12 weeks of therapy significantly reduced febrile episodes and deaths without increasing healthcare associated infections or carriage. Trial Registration Number: ISRCTN number: ISRCTN51731976 and EudraCT number: 2011-000366-35. Funding Statement: Funded by NIHR Health Technology Assessment Programme. 08/116/69. Declaration of Interests: None. Ethics Approval Statement: Approved by the UK Coventry & Warwickshire Multi-Research Ethics Committee on 29th July 2011. All patients provided written informed consent.

Sequencing of the bla IMP-4 -carrying C. freundii B38 using the PacBio SMRT technique revealed that the genome contained a chromosome of 5,134,500 bp and three plasmids, pOZ172 (127,005 bp), pOZ181 (277,592 bp), and pOZ182 (18,467 bp). Plasmid pOZ172 was identified as IncFIIY, like pP10164-NDM and pNDM-EcGN174. It carries a class 1 integron with four cassettes ( bla IMP-4 - qacG2 - aacA4 - aphA15 ) and a complete hybrid tni module ( tniR - tniQ - tniB - tniA ). The recombination of tniR from Tn 402 (identical) with tniQBA from Tn 5053 (99%) occurred within the res site of Tn 402 / 5053 . The Tn 402/5053 -like integron, named Tn 6017 , was inserted into Tn 1722 at the res II site. The replication, partitioning, and transfer systems of pOZ181 were similar to those of IncHI2 plasmids (e.g., R478) and contained a sul1 -type class 1 integron with the cassette array orf-dfrA1-orf-gcu37-aadA5 linked to an upstream Tn 1696 tnpA-tnpR and to a downstream 3′ conserved sequence (3′-CS) and IS CR1 . A Tn 2 transposon encoding a bla TEM-1 β-lactamase was identified on pOZ182. Other interesting resistance determinants encoded on the B38 chromosome included multidrug resistance (MDR) efflux pumps, an AmpC β-lactamase, and resistances to Cu, Ag, As, and Zn. This is the first report of a complete tni module linked to a bla IMP-4 -carrying class 1 integron, which, together with other recently reported non- sul1 integrons, represents the emergence of a distinct evolutionary lineage of class 1 integrons lacking a 3′-CS ( qacEΔ1 - sul1 ). The unique cassette array, complete tni module of Tn 6017 , and incompatibility group of pOZ172 suggest a bla IMP-4 evolutionary pathway in C. freundii B38 different from that for other bla IMP-4 genes found in Gram-negative bacteria in the Western Pacific region.

Summary Faecal samples from 1365 healthy asymptomatic volunteers from four regions in England were screened for the presence of Clostridium difficile between December 2013 and July 2014. The carriage rate of C. difficile in healthy patients was 0.5%, which is lower than reported previously. This study demonstrates that the true community reservoir of C. difficile in the healthy UK population is very low and is, therefore, unlikely to be a reservoir for infections diagnosed in the hospital setting.

Background Faecal Microbiota Transplant (FMT) has improved outcomes for the treatment of Clostridioides difficile infection (CDI) compared to antibiotic therapy. FMT is classified as a medicinal product in the United Kingdom, similar to the USA and Canada, limiting supply via stool banks without appropriate licencing. In the largest UK cohort to date, we describe the clinical outcomes for 124 patients receiving FMT for recurrent or refractory CDI and present a framework to produce FMT as a licenced medicinal product. Methods Anonymous unrelated healthy donors, screened via health assessment and microbiological testing donated stool. In aerobic conditions FMT aliquots were prepared for immediate use or frozen storage, following a production framework developed to comply with Good Manufacturing Practice. Outcome measures were clinical response to FMT defined as resolution of diarrhoea within seven days and clinical cure defined as response without diarrhoea recurrence at 90 days. Findings Clinical response was 83·9% (95% CI 76·0%–90·0%) after one treatment. Clinical cure was 78·2% (95% CI 67·4%–89·0%) across the cohort. Refractory cases appeared to have a lower initial clinical response rate compared to recurrent cases, however at day 90 there were no differences observed between these groups. Interpretation The methodology developed here enabled successful licencing of FMT by The Medicines and Healthcare products Regulatory Agency as a medicinal product. This has widened the availability of FMT in the National Health Service via a stool bank and can be applied in other centres across the world to improve access to safe and quality assured treatments.

In this review, we highlight the variations of gut resistome studies, which may preclude comparisons and translational interpretations. Of 22 included studies, a range of 12 to 2000 antibiotic resistance (AR) genes were profiled. Overall, studies defined a healthy gut resistome as subjects who had not taken antibiotics in the last three to 12 months prior to sampling. In studies with de novo assembly, AR genes were identified based on variable nucleotide or amino acid sequence similarities. Different marker genes were used for defining resistance to a given antibiotic class. Validation of phenotypic resistance in the laboratory is frequently lacking. Cryptic resistance, collateral sensitivity and the interaction with repressors or promotors were not investigated. International consensus is needed for selecting marker genes to define resistance to a given antibiotic class in addition to uniformity in phenotypic validation and bioinformatics pipelines.

The widespread use of antibacterial drugs over the last 70 years has brought immense benefits to human health at the price of increasing drug inefficacy. Antibacterial agents have a strong selective effect in both favouring resistant strains and allowing particular species and families of bacteria to prosper, especially in the healthcare setting. Whilst important Gram-positive bacterial pathogens such as Staphylococcus aureus and Streptococcus pneumoniae caused concern over the last 20 years because of the spread of antibiotic-resistant strains, Enterobacteriaceae have become the biggest challenge. They have very efficient mechanisms for genetic exchange, as illustrated by the emergence and rapid spread of CTX-M β-lactamases and the carbapenemases. The unique epidemiology of Enterobacteriaceae, with substantial numbers colonizing the mammalian gut and subsequent release into and spread in the environment, presents a significant threat to human health because of the high levels of exposure for the whole community. The use of antimicrobials in agriculture combined with global movements of people, animals and food, arising from worldwide industrialization, generates a diversity and level of resistance not seen previously. Control will require globally coordinated interventions similar to those needed to ameliorate climate change.

Interest in the therapeutic potential of faecal microbiota transplant (FMT) has been increasing globally in recent years, particularly as a result of randomised studies in which it has been used as an intervention. The main focus of these studies has been the treatment of recurrent or refractoryClostridium difficileinfection (CDI), but there is also an emerging evidence base regarding potential applications in non-CDI settings. The key clinical stakeholders for the provision and governance of FMT services in the UK have tended to be in two major specialty areas: gastroenterology and microbiology/infectious diseases. While the National Institute for Health and Care Excellence (NICE) guidance (2014) for use of FMT for recurrent or refractory CDI has become accepted in the UK, clear evidence-based UK guidelines for FMT have been lacking. This resulted in discussions between the British Society of Gastroenterology (BSG) and Healthcare Infection Society (HIS), and a joint BSG/HIS FMT working group was established. This guideline document is the culmination of that joint dialogue.

Interest in the therapeutic potential of faecal microbiota transplant (FMT) has been increasing globally in recent years, particularly as a result of randomised studies in which it has been used as an intervention. The main focus of these studies has been the treatment of recurrent or refractory

The Working Party makes more than 100 tabulated recommendations in antimicrobial prescribing for the treatment of infections caused by multidrug-resistant (MDR) Gram-negative bacteria (GNB) and suggest further research, and algorithms for hospital and community antimicrobial usage in urinary infection. The international definition of MDR is complex, unsatisfactory and hinders the setting and monitoring of improvement programmes. We give a new definition of multiresistance. The background information on the mechanisms, global spread and UK prevalence of antibiotic prescribing and resistance has been systematically reviewed. The treatment options available in hospitals using intravenous antibiotics and in primary care using oral agents have been reviewed, ending with a consideration of antibiotic stewardship and recommendations. The guidance has been derived from current peer-reviewed publications and expert opinion with open consultation. Methods for systematic review were NICE compliant and in accordance with the SIGN 50 Handbook; critical appraisal was applied using AGREE II. Published guidelines were used as part of the evidence base and to support expert consensus. The guidance includes recommendations for stakeholders (including prescribers) and antibiotic-specific recommendations. The clinical efficacy of different agents is critically reviewed. We found there are very few good-quality comparative randomized clinical trials to support treatment regimens, particularly for licensed older agents. Susceptibility testing of MDR GNB causing infection to guide treatment needs critical enhancements. Meropenem- or imipenem-resistant Enterobacteriaceae should have their carbapenem MICs tested urgently, and any carbapenemase class should be identified: mandatory reporting of these isolates from all anatomical sites and specimens would improve risk assessments. Broth microdilution methods should be adopted for colistin susceptibility testing. Antimicrobial stewardship programmes should be instituted in all care settings, based on resistance rates and audit of compliance with guidelines, but should be augmented by improved surveillance of outcome in Gram-negative bacteraemia, and feedback to prescribers. Local and national surveillance of antibiotic use, resistance and outcomes should be supported and antibiotic prescribing guidelines should be informed by these data. The diagnosis and treatment of both presumptive and confirmed cases of infection by GNB should be improved. This guidance, with infection control to arrest increases in MDR, should be used to improve the outcome of infections with such strains. Anticipated users include medical, scientific, nursing, antimicrobial pharmacy and paramedical staff where they can be adapted for local use.

Global trade and mobility of people has increased rapidly over the last 20 years. This has had profound consequences for the evolution and the movement of antibiotic resistance genes. There is increasing exposure of populations all around the world to resistant bacteria arising in the emerging economies. Arguably the most important development of the last two decades in the field of antibiotic resistance is the emergence and spread of extended-spectrum β-lactamases (ESBLs) of the CTX-M group. A consequence of the very high rates of ESBL production among Enterobacteriaceae in Asian countries is that there is a substantial use of carbapenem antibiotics, resulting in the emergence of plasmid-mediated resistance to carbapenems. This article reviews the emergence and spread of multidrug-resistant Gram-negative bacteria, focuses on three particular carbapenemases--imipenem carbapenemases, Klebsiella pneumoniae carbapenemase, and New Delhi metallo-β-lactamase--and highlights the importance of control of antibiotic use.

Objectives To retrospectively analyse routine susceptibility testing data to describe antimicrobial non-susceptibility trends in isolates of Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa from urine samples in a population of 5.6 million people over a 4 year period. Methods De-duplicated laboratory data submitted to the AmSurv surveillance system from the West Midlands region of England and results of submissions to the Antimicrobial Resistance and Healthcare Associated Infections Reference Unit were extracted for the period 2010–13. Descriptive analysis of the non-susceptibility of selected Gram-negative organisms to key antibiotics, as recommended for monitoring in the UK Five Year Antimicrobial Resistance Strategy, was undertaken. Results During the study period, there were 431 461 reports for E. coli, 23 786 for K. pneumoniae and 6985 for P. aeruginosa from urine specimens. These represented 61%, 3% and 1%, respectively, of all organisms isolated from urine specimens. There was a linear increase in non-susceptibility to third-generation cephalosporins for E. coli and K. pneumoniae, and to ciprofloxacin for E. coli, in specimens from both hospital and community settings (P Conclusions Automated antimicrobial resistance surveillance enabled, for the first time in England, the systematic monitoring of resistance in bacteria responsible for urinary tract infections in a defined population, and thereby provided a representative indication of the burden of resistance in Gram-negative bacteria in hospital and community settings.

We analyzed China’s current use of and microbial resistance to antibiotics, and possible means of reducing antimicrobial resistance. Interventions like executive orders within clinical settings and educational approach with vertical approaches rather than an integrated strategy to curb the use of antimicrobials remain limited. An underlying problem is the system of incentives that has resulted in the intensification of inappropriate use by health professionals and patients. There is an urgent need to explore the relationship between financial and non-financial incentives for providers and patients, to eliminate inappropriate incentives. China’s national health reforms have created an opportunity to contain inappropriate use of antibiotics through more comprehensive and integrated strategies. Containment of microbial resistance may be achieved by strengthening surveillance at national, regional and hospital levels; eliminating detrimental incentives within the health system; and changing prescribing behaviors to a wider health systems approach, to achieve long-term, equitable and sustainable results and coordinate stakeholders’ actions through transparent sharing of information.

Hospital acquired infection is a major cause of morbidity and mortality and regimes to prevent infection are crucial in infection control. These include decolonisation of at-risk patients of carriage of MRSA which is commonly achieved by protocols that include the use of chlorhexidine, or octenidine as biocidal agents. There is however no standardised single decolonisation regime agreed upon in the UK or other countries and protocols include a variety of active agents. Antibiotic resistant bacteria cause major problems in hospital medicine and concern has been raised regarding the development of biocide resistance which would cause decolonisation regimes to become unreliable. In this study, we assembled a panel of isolates of S. aureus including isolates collected before the development of chlorhexidine and octenidine through to a contemporaneous panel of isolates from a major hospital trust in the UK during a period when the decolonisation regime was altered. We observed significant increases in the MIC and MBC of chlorhexidine in isolates collected from periods of high usage of chlorhexidine. No isolates had a significantly altered MIC or MBC of octenidine apart from those collected after octenidine was introduced into the trust where isolates with four-fold decreases in susceptibility emerged. There was no suggestion of cross-resistance between the two biocidal agents. A combination of VNTR, PCR for qac genes and whole genome sequencing was used to type isolates and examine possible mechanisms of resistance. The typing data showed no expansion of a single strain was associated with decreased biocide tolerance and isolates with increased chlorhexidine MIC and MBCs were found from different clonal complexes; CC8, CC22 and CC30. Biocide susceptibility did not correlate with carriage of qac efflux pump genes – carriage of qacA and qacB was detected but, with one exception was restricted to isolates of CC8. Analysis of genome sequence data for closely related pairs of strains with differential biocide susceptibility revealed no common mutations or carriage of accessory elements that correlated with biocide tolerance. Mutations with the NorA or NorB efflux pumps, previously associated with chlorhexidine export were identified suggesting this may be an important mechanism of biocide tolerance. The clinical relevance of decreased biocide tolerance in terms of efficacy of decolonisation therapies remains to be established but we present evidence here that isolates are evolving in the face of biocide challenge in patients and that changes to decolonisation regimes are reflected in changes in susceptibility of isolates. More work is needed to assess the impact of these changes to ensure effective and robust decolonisation protocols remain in place.

Objectives To assess the effect of general practice characteristics and antibiotic prescribing on the number of non-susceptible Escherichia coli isolated from urine specimens submitted from community settings, we undertook an ecological study of the general practice population in the West Midlands. Methods Descriptive analysis and multilevel modelling of temporal trends in antibiotic prescribing and non-susceptibility of E. coli urine isolates to a range of antibiotics prescribed in the community over a 4 year period. Results Nine of the 16 antibiotic prescribing/non-susceptibility combinations demonstrated a significant statistical linear correlation with non-susceptibility either for prescribing in a quarter or for prescribing within the previous 12 months. The magnitude of the effect varied, from a 0.3% increase in the odds of non-susceptibility to ampicillin/amoxicillin (when prescribing ampicillin/amoxicillin) to a 6.3% increase in the odds of non-susceptibility to nitrofurantoin (when prescribing nitrofurantoin) for an increase of 50 DDDs per 1000 practice population within a quarter (equivalent to ∼10 courses of antibiotics). In all 16 models, single-handed general practices were shown to have a significant association with increased numbers of non-susceptible E. coli urine isolates (adjusted ORs 1.083-1.657). Increased prescribing of ampicillin/amoxicillin in winter periods was associated with increased non-susceptibility of E. coli isolated from urine specimens. Conclusions Small increases in antibiotic prescribing in individual general practices reduce the number of susceptible bacteria in the practice population. To maintain the effectiveness of available treatment, antibiotic stewardship should be encouraged and supported within each practice.

Antibiotic-resistant bacteria (ARB) present a global public health problem. With numbers of community-acquired resistant infections increasing, understanding the mechanisms by which people are exposed to and colonised by ARB can help inform effective strategies to prevent their spread. The role natural environments play in this is poorly understood. This is the first study to combine surveillance of ARB in bathing waters, human exposure estimates and association between exposure and colonisation by ARB in water users.97 bathing water samples from England and Wales were analysed for the proportion of E. coli harbouring bla11 of 97 bathing waters sampled were found to contain blaSurfers are at risk of exposure to and colonisation by clinically important antibiotic-resistant E. coli in coastal waters. Further research must be done on the role natural environments play in the transmission of ARB.

Pneumococcal disease has a high burden in adults in the United Kingdom (UK); however, the total burden is underestimated, principally because most cases of community-acquired pneumonia (CAP) are non-invasive. Research into pneumonia receives poor funding relative to its disease burden (global mortality, disability-adjusted life years, and years lived with disability), ranking just 20 out of 25 for investment in infectious diseases in the UK. The current accuracy of data for establishing incidence rates is questionable, and it is a reflection of the paucity of research that much of the background information available derives from nearly 30 years ago. Given the relationship between CAP and mortality (pneumonia accounts for 29,000 deaths per annum in the UK, and 5–15% of patients hospitalised with CAP die within 30 days of admission), and the increasing threat of antimicrobial resistance associated with inappropriate antibiotic prescribing, such neglect of a highly prevalent problem is concerning. In this Call to Action, we explore the poorly understood burden of CAP in the UK, discuss the importance of an accurate diagnosis and appropriate treatment, and suggest how national collaboration could improve the management of an often life-threatening, yet potentially preventable disease.

The diarrheal pathogen Clostridium difficile consists of at least six distinct evolutionary lineages. The RT017 lineage is anomalous, as strains only express toxin B, compared to strains from other lineages that produce toxins A and B and, occasionally, binary toxin. Historically, RT017 initially was reported in Asia but now has been reported worldwide. We used whole-genome sequencing and phylogenetic analysis to investigate the patterns of global spread and population structure of 277 RT017 isolates from animal and human origins from six continents, isolated between 1990 and 2013. We reveal two distinct evenly split sublineages (SL1 and SL2) of C. difficile RT017 that contain multiple independent clonal expansions. All 24 animal isolates were contained within SL1 along with human isolates, suggesting potential transmission between animals and humans. Genetic analyses revealed an overrepresentation of antibiotic resistance genes. Phylogeographic analyses show a North American origin for RT017, as has been found for the recently emerged epidemic RT027 lineage. Despite having only one toxin, RT017 strains have evolved in parallel from at least two independent sources and can readily transmit between continents.

Anthropogenic inputs increase levels of antimicrobial resistance (AMR) in the environment, however, it is unknown how these inputs create this observed increase, and if anthropogenic sources impact AMR in environmental bacteria. The aim of this study was to characterise the role of waste water treatment plants (WWTPs) in the dissemination of class 1 integrons (CL1s) in the riverine environment. Using sample sites from upstream and downstream of a WWTP, we demonstrate through isolation and culture-independent analysis that WWTP effluent significantly increases both CL1 abundance and antibiotic resistance in the riverine environment. Characterisation of CL1-bearing isolates revealed that CL1s were distributed across a diverse range of bacteria, with identical complex genetic resistance determinants isolated from both human-associated and common environmental bacteria across connected sites. Over half of sequenced CL1s lacked the 3'-conserved sequence ('atypical' CL1s); surprisingly, bacteria carrying atypical CL1s were on average resistant to more antibiotics than bacteria carrying 3'-CS CL1s. Quaternary ammonium compound (QAC) resistance genes were observed across 75% of sequenced CL1 gene cassette arrays. Chemical data analysis indicated high levels of boron (a detergent marker) downstream of the WWTP. Subsequent phenotypic screening of CL1-bearing isolates demonstrated that ~90% were resistant to QAC detergents, with in vitro experiments demonstrating that QACs could solely select for the transfer of clinical antibiotic resistance genes to a naive Escherichia coli recipient. In conclusion, this study highlights the significant impact of WWTPs on environmental AMR, and demonstrates the widespread carriage of clinically important resistance determinants by environmentally associated bacteria.

The environment harbours a significant diversity of uncultured bacteria and a potential source of novel and extant resistance genes which may recombine with clinically important bacteria disseminated into environmental reservoirs. There is evidence that pollution can select for resistance due to the aggregation of adaptive genes on mobile elements. The aim of this study was to establish the impact of waste water treatment plant (WWTP) effluent disposal to a river by using culture independent methods to study diversity of resistance genes downstream of the WWTP in comparison to upstream. Metagenomic libraries were constructed in Escherichia coli and screened for phenotypic resistance to amikacin, gentamicin, neomycin, ampicillin and ciprofloxacin. Resistance genes were identified by using transposon mutagenesis. A significant increase downstream of the WWTP was observed in the number of phenotypic resistant clones recovered in metagenomic libraries. Common β-lactamases such as blaTEM were recovered as well as a diverse range of acetyltransferases and unusual transporter genes, with evidence for newly emerging resistance mechanisms. The similarities of the predicted proteins to known sequences suggested origins of genes from a very diverse range of bacteria. The study suggests that waste water disposal increases the reservoir of resistance mechanisms in the environment either by addition of resistance genes or by input of agents selective for resistant phenotypes.

IntroductionImbalance of the gut microbiome is key to the pathogenesis of ulcerative colitis (UC). Faecal microbiota transplant (FMT) is the transfer of homogenised and filtered faeces from a healthy individual to the gastrointestinal tract of a patient with disease. Published datasets show a positive signal for the use of FMT to treat UC, but the optimal route and dose of FMT remain unanswered.Methods and analysisThis prospective, multi-centre open-label, randomised pilot study will assess two possible routes of FMT delivery, via the nasogastric (NG) route or by delivery to the COLON, in 30 patients with active UC recruited from three sites in the UK. Stool will be collected from healthy screened donors, processed, frozen and stored under a Medicines and Healthcare products Regulatory Agency (MHRA) “specials” manufacturing licence held at the University of Birmingham Microbiome Treatment Centre. Thawed FMT samples will be administered to patients either via eight nasogastric infusions given initially over 4 days starting on the day of randomisation, and then again for 4 days in week 4 for foregut delivery (total of 240 g of stool) or via one colonoscopic infusion followed by seven weekly enemas according to the hindgut protocol (total of 360 g of stool). Patients will be followed up weekly for 8 weeks, and then at 12 weeks. The aims of this pilot study are (1) to determine which FMT administration route (NG or COLON) should be investigated in a randomised double-blind, placebo-controlled trial and (2) to determine if a full randomised controlled trial is feasible. The primary outcome will be a composite assessment of both qualitative and quantitative data based on efficacy (clinical response), acceptability and safety. At the end of the pilot study, decisions will be made regarding the feasibility of a full randomised double-blind, placebo-controlled trial and, if deemed feasible, which route of administration should be used in such a study.Ethics and disseminationEthical approval for this study has been obtained from the East Midlands-Nottingham Research Ethics Committee (REC 17/EM/0274). At the end of the study, findings will be reported at national and international gastroenterology meetings and published in peer-reviewed journals.Trial registration numberNCT74072945

Infections with >1 Mycobacterium tuberculosis strain(s) are underrecognized. We show, in vitro and in vivo, how first-line treatment conferred a competitive growth advantage to amplify a multidrug-resistant M. tuberculosis strain in a patient with mixed infection. Diagnostic techniques that identify mixed tubercle bacilli populations are needed to curb the spread of multidrug resistance.

During the past 10 years, multidrug-resistant Gram-negative Enterobacteriaceae have become a substantial challenge to infection control. It has been suggested by clinicians that the effectiveness of antibiotics is in such rapid decline that, depending on the pathogen concerned, their future utility can be measured in decades or even years. Unless the rise in antibiotic resistance can be reversed, we can expect to see a substantial rise in incurable infection and fatality in both developed and developing regions. Antibiotic resistance develops through complex interactions, with resistance arising by de-novo mutation under clinical antibiotic selection or frequently by acquisition of mobile genes that have evolved over time in bacteria in the environment. The reservoir of resistance genes in the environment is due to a mix of naturally occurring resistance and those present in animal and human waste and the selective effects of pollutants, which can co-select for mobile genetic elements carrying multiple resistant genes. Less attention has been given to how anthropogenic activity might be causing evolution of antibiotic resistance in the environment. Although the economics of the pharmaceutical industry continue to restrict investment in novel biomedical responses, action must be taken to avoid the conjunction of factors that promote evolution and spread of antibiotic resistance.

SETTING Birmingham, United Kingdom, 2010-2014. OBJECTIVE To investigate predictors for clustering of tuberculosis (TB) cases and cluster size and to evaluate the impact of cluster investigation using social network data. DESIGN Retrospective observational cohort study. Prioritised cases linked using 24-locus mycobacterial interspersed repetitive units-variable number of tandem repeats (MIRU-VNTR) were interviewed using a social network approach to find epidemiological links. RESULTS Of 2055 TB cases notified, 56% could be typed. Clustering was associated with younger age, UK birth, Black Caribbean ethnicity, social risk factors, pulmonary TB and negative human immunodeficiency virus status. Only UK birth and presence of more than one social risk factor were associated with larger cluster size, while drug resistance was associated with smaller cluster size. Social network data from 139/431 clustered cases found new epidemiological links in 11/19 clusters with ⩾5 members (undirected median network density 0.09, interquartile range 0.05-0.4). Ninety-eight additional contacts were assessed, with one case of active TB and 24 with latent tuberculous infection diagnosed. CONCLUSION A social network approach increased knowledge of likely transmission events, but few additional TB cases were diagnosed. Obtaining social network data for all typed and untyped TB cases may improve contact tracing and reduce unexpected transmission detected from molecular data.

The early identification of outbreaks is crucial for the control of Clostridium difficile infection. This study aimed to determine if the number of hospital-acquired C. difficile infections could be reduced by rapidly typing C. difficile strains using multiple-locus variable-number tandem-repeat analysis (MLVA) compared to typing using PCR ribotyping. A total of 16 hospitals were recruited to the study, and all periods of increased incidence (PIIs) of C. difficile infection were identified. The hospitals were randomized into two study arms, the test and the control, with all isolates typed in the test using MLVA and in the control using PCR ribotyping. Following a PII, each hospital received a structured questionnaire regarding control measures implemented or stopped prior to or following the typing results. During the study period, there were a total of 1,682 hospital-apportioned C. difficile toxin-positive cases, with 868 in the control and 814 in the test, with modeling demonstrating no differences between the two arms. A total of 245 PIIs occurred, involving 785 patients. There was a significant difference in the mean turnaround time between the ribotyping and MLVA typing (13.6 and 5.3 days, respectively [ P < 0.001]). The discriminatory ability of MLVA was greater than ribotyping, with 85 outbreaks being confirmed by ribotyping and 62 by MLVA. In the test arm, 40.6% of respondents strongly agreed that the typing result had aided their management of clusters, as opposed to 9.9% in the control. The study demonstrated the utility of rapidly typing C. difficile strains, demonstrating that it aided the management of clusters, enabling effective targeting of infection control resources.

BACKGROUND: Marked increases in Clostridium difficile infection (CDI) incidence, driven by epidemic strain spread, is a global phenomenon. METHODS: The Clostridium difficile Ribotyping Network (CDRN) was established in 2007 as part of enhanced CDI surveillance in England, to facilitate the recognition and control of epidemic strains. We report on changes in CDI epidemiology in England in the first 3 years of CDRN. RESULTS: CDRN received 12,603 fecal specimens, comprising significantly (P < .05) increasing numbers and proportions of national CDI cases in 2007-2008 (n = 2109, 3.8%), 2008-2009 (n = 4774, 13.2%), and 2009-2010 (n = 5720, 22.3%). The C. difficile recovery rate was 90%, yielding 11,294 isolates for ribotyping. Rates of 9 of the 10 most common ribotypes changed significantly (P < .05) during 2007-2010. Clostridium difficile ribotype 027 predominated, but decreased markedly from 55% to 36% and 21% in 2007-2008, 2008-2009, and 2009-2010, respectively. The largest regional variations in prevalence occurred for ribotypes 027, 002, 015, and 078. Cephalosporin and fluoroquinolone use in CDI cases was reported significantly (P < .05) less frequently during 2007-2010. Mortality data were subject to potential reporting bias, but there was a significant decrease in CDI-associated deaths during 2007-2010, which may have been due to multiple factors, including reduced prevalence of ribotype 027. CONCLUSIONS: Access to C. difficile ribotyping was associated with significant changes in the prevalence of epidemic strains, especially ribotype 027. These changes coincided with markedly reduced CDI incidence and related mortality in England. CDI control programs should include prospective access to C. difficile typing and analysis of risk factors for CDI and outcomes.

Antibiotics are widely used in human and veterinary medicine for the prevention and treatment of infectious diseases. This practice has led to the emergence of antimicrobial-resistant bacteria in both humans and animals. The potential role that animals, particularly livestock, might play as potential reservoirs of antibiotic resistance genes has been recognized, and it is currently a cause of public health concern. The impact of animal and human antibiotic usage on the emergence and persistence of resistant bacteria and the precise transfer pathways for resistance genes between humans and animals are not currently fully understood. As part of the remit of the UK Advisory Committee on Antimicrobial Resistance and Healthcare-Associated Infection (ARHAI), two main areas were addressed, namely methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Gram-negative bacteria, where both the human and veterinary health sectors share interests. We review the current knowledge of MRSA and resistant Gram-negative bacteria, and provide guidance on occupational risks for veterinary healthcare workers relating to animals infected or colonized with MRSA. Findings and recommendations for further work across disciplines and future research in multidrug-resistant Gram-negative bacteria are also presented. Working collaboratively across disciplines is essential in order to better understand and challenge an important human and animal health problem: antimicrobial resistance.

OBJECTIVES To characterize plasmids encoding extended-spectrum β-lactamases (ESBLs) from a recent UK collection of clinical Escherichia coli isolates. METHODS The isolates comprised 118 ESBL producers referred from 54 laboratories. Plasmids were transferred by electroporation, and their incompatibility groups, associated addiction systems and resistance genes with the flanking genetic environments were identified by PCR or sequencing. RESULTS Seventy isolates had plasmids encoding CTX-M-15 (n = 53), CTX-M-14 (n =9), CTX-M-27 (n = 1), CTX-M-3 (n = 2) and SHV-12 (n = 5) ESBLs that were transformable; non-transformable ESBLs were mainly CTX-M enzymes (42/48). Most transformable bla(CTX-M-15) genes (43/53) were harboured on single replicon or multireplicon IncF plasmids, with IncFIA4-FIB1-FII31 (n = 11) and IncFIA1-FII2 (n = 15) being most frequent; the latter included eight pEK499 plasmids, typical of UK epidemic strain A. Plasmids harbouring bla(CTX-M-14) belonged variously to IncF, IncI1 and IncHI2 types, and 16 encoding CTX-M or SHV enzymes were non-typeable. Only IncF plasmid types carried the addiction systems sought and those with bla(CTX-M-15) frequently harboured bla(OXA-1) and aac(6')-Ib-cr, and often transferred trimethoprim and tetracycline resistance; those with bla(CTX-M-14) encoded trimethoprim, sulphonamide, streptomycin and tetracycline resistance. Most ESBL genes were associated with the well-known mobile elements ISEcp1 and IS26, but nearly half (23/55) of the ISEcp1 sequences upstream of bla(CTX-M-15) were interrupted by an IS26 at various positions. CONCLUSIONS Most ESBLs (70/118) were encoded by transformable plasmids, although a sizable minority could not be transformed. The majority of transformable plasmids (51/70; 72.9%) were diverse multiresistant IncF types possessing multiple addiction systems. The spread of bla(CTX-M-15) can be attributed not just to clonal expansion, but also to the horizontal dissemination of related plasmids.

PCR ribotyping is currently used in many countries for epidemiological investigation to track transmission and to identify emerging variants of Clostridium difficile . Although PCR ribotyping differentiates over 300 types, it is not always sufficiently discriminatory for epidemiological investigations particularly for common ribotypes, e.g., ribotypes 027, 106, and 017. Multilocus variable-number tandem-repeat analysis (MLVA) is a highly discriminatory molecular subtyping method that has been applied to a number of bacterial species for high-level subtyping. Two MLVA typing schemes for C. difficile have been previously published, each utilizing seven variable-number tandem-repeat (VNTR) loci on the genome with four loci common to both schemes. Although these schemes are good genotyping methods with the ability to discriminate between isolates, they do not identify the ribotype. We show here that increasing the number of VNTR loci to 15, creating the extended MLVA (eMLVA) scheme, we have successfully subtyped all clinically significant ribotypes while still clustering isolates in concordance with PCR ribotyping. The eMLVA scheme developed here provides insight into the genetic diversity of the C. difficile population at both global and cross-infection clusters in patient levels, with the possibility of replacing PCR ribotyping.

Due to an increase in bovine tuberculosis in cattle in the United Kingdom, we investigated the characteristics of Mycobacterium bovis infection in humans and assessed whether extensive transmission of M. bovis between humans has occurred. A cross-sectional study linking demographic, clinical, and DNA fingerprinting (using 15-locus mycobacterial interspersed repetitive-unit–variable-number tandem-repeat [MIRU-VNTR] typing) data on cases reported between 2005 and 2008 was undertaken. A total of 129 cases of M. bovis infection in humans were reported over the period, with a decrease in annual incidence from 0.065 to 0.047 cases per 100,000 persons. Most patients were born pre-1960, before widespread pasteurization was introduced (73%), were of white ethnicity (83%), and were born in the United Kingdom (76%). A total of 102 patients (79%) had MIRU-VNTR typing data. A total of 31 of 69 complete MIRU-VNTR profiles formed eight distinct clusters. The overall clustering proportion determined using the n − 1 method was 33%. The largest cluster, comprising 12 cases, was indistinguishable from a previously reported West Midlands outbreak strain cluster and included those cases. This cluster was heterogeneous, having characteristics supporting recent zoonotic and human-to-human transmission as well as reactivation of latent disease. Seven other, smaller clusters identified had demographics supporting recrudescence rather than recent infection. A total of 33 patients had incomplete MIRU-VNTR profiles, of which 11 may have yielded 2 to 6 further small clusters if typed to completion. The incidence of M. bovis in humans in the United Kingdom remains low, and the epidemiology is predominantly that of reactivated disease.

The impact of human activity on the selection for antibiotic resistance in the environment is largely unknown, although considerable amounts of antibiotics are introduced through domestic wastewater and farm animal waste. Selection for resistance may occur by exposure to antibiotic residues or by co-selection for mobile genetic elements (MGEs) which carry genes of varying activity. Class 1 integrons are genetic elements that carry antibiotic and quaternary ammonium compound (QAC) resistance genes that confer resistance to detergents and biocides. This study aimed to investigate the prevalence and diversity of class 1 integron and integron-associated QAC resistance genes in bacteria associated with industrial waste, sewage sludge and pig slurry. We show that prevalence of class 1 integrons is higher in bacteria exposed to detergents and/or antibiotic residues, specifically in sewage sludge and pig slurry compared with agricultural soils to which these waste products are amended. We also show that QAC resistance genes are more prevalent in the presence of detergents. Studies of class 1 integron prevalence in sewage sludge amended soil showed measurable differences compared with controls. Insertion sequence elements were discovered in integrons from QAC contaminated sediment, acting as powerful promoters likely to upregulate cassette gene expression. On the basis of this data, >1 × 1019 bacteria carrying class 1 integrons enter the United Kingdom environment by disposal of sewage sludge each year.