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Background: Exposure to antibiotics predisposes to dysbiosis and Clostridioides difficile infection (CDI) that can be severe, recurrent (rCDI), and life-threatening. Nonselective drugs that treat CDI and perpetuate dysbiosis are associated with rCDI, in part due to loss of microbiome-derived secondary bile acid (SBA) production. Ridinilazole is a highly selective drug designed to treat CDI and prevent rCDI., Methods: In this phase 3 superiority trial, adults with CDI, confirmed with a stool toxin test, were randomized to receive 10 days of ridinilazole (200 mg twice daily) or vancomycin (125 mg 4 times daily). The primary endpoint was sustained clinical response (SCR), defined as clinical response and no rCDI through 30 days after end of treatment. Secondary endpoints included rCDI and change in relative abundance of SBAs., Results: Ridinilazole and vancomycin achieved an SCR rate of 73% versus 70.7%, respectively, a treatment difference of 2.2% (95% CI: -4.2%, 8.6%). Ridinilazole resulted in a 53% reduction in recurrence compared with vancomycin (8.1% vs 17.3%; 95% CI: -14.1%, -4.5%; P = .0002). Subgroup analyses revealed consistent ridinilazole benefit for reduction in rCDI across subgroups. Ridinilazole preserved microbiota diversity, increased SBAs, and did not increase the resistome. Conversely, vancomycin worsened CDI-associated dysbiosis, decreased SBAs, increased Proteobacteria abundance (∼3.5-fold), and increased the resistome., Conclusions: Although ridinilazole did not meet superiority in SCR, ridinilazole greatly reduced rCDI and preserved microbiome diversity and SBAs compared with vancomycin. These findings suggest that treatment of CDI with ridinilazole results in an earlier recovery of gut microbiome health. Clinical Trials Registration.Ri-CoDIFy 1 and 2: NCT03595553 and NCT03595566., Competing Interests: Potential conflicts of interest. K. W. G. received grants paid to his institution from Acurx, Paratek, Cidara, Therapeutics, and Seres Health. P. C. O. received faculty grant/research support from Merck, Sharp and Dohme, Deinove Pharmaceuticals, Summit Pharmaceuticals, Melinta Pharmaceuticals, the National Institutes of Health (NIH) National Institute of Allergy and Infectious Diseases (NIAID), Kieberg Foundation, and Napo Pharmaceutical; is a consultant to Napo Pharmaceutical, Ferring Pharmaceutical, Summit Pharmaceutical, SNIPR Biome Company, Haleon, and SNIPR; received payment for speaking from Colegio Mexicano de Medicina Interna; participated on a Scientific Advisory Board for Napo Pharmaceuticals; and has stock or stock options from Moderna, Pfizer, Haleon, Biontech, GSK, Novavax, AstraZeneca, Beam, and Johnson & Johnson. T. L. attended advisory board meetings for Summit, Inc, and received payments for the conduct of clinical trials for Seres Therapeutics, Finch Therapeutics, and Rebiotix/Ferring. E. D. and D. J. were employees of Summit, Inc, and owned stock options in Summit Therapeutics (parent company) at the time of manuscript submission. J. G. M. is a part-time employee and owns stock options in Summit, Inc. E. R. D. has received grants paid to his institution from Theriva Biologics and Ferring and consulting fees from Pfizer, Merck, Seres, Abbott, AstraZeneca, Summit, GSK, and Ferring. M. W. has received consulting fees from AiCuris, Bayer, Crestone, Da Volterra, Deinove, EnteroBiotix, The European Tissue Symposium, Ferring, GSK, Menarini, Merck, Nestlé, Paion, Paratek, Pfizer, Phico Therapeutics, Qpex Biopharma, Seres, Surface Skins, Summit, Tillotts, Vaxxilon/Idorsia, and Vedanta; lecture fees from GSK, Merck, Pfizer, Seres, and Tillotts; and grant support from Almirall, Da Volterra, EnteroBiotix, GSK, Merck, MicroPharm, Nabriva, Paratek, Pfizer, Seres, Summit, The European Tissue Symposium, and Tillotts. J. L., B. Y. C., J. S., L. S., and F. C. are employees of Summit, Inc, and own Summit stock options. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2024. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Background: Fidaxomicin is a first-line treatment for Clostridioides difficile infections (CDIs). Fidaxomicin resistance has rarely been reported in this urgent antimicrobial resistance threat as defined by the CDC., Objectives: To report a case of fidaxomicin-resistant C. difficile isolation in a patient treated by fidaxomicin, characterize the genetic determinant for resistance and the consequences on pathophysiological traits, and review the literature., Patient and Methods: A 38-year-old male patient with several risk factors for CDI experienced three episodes of hospital-acquired CDI and received fidaxomicin for the first episode. The successive isolates were subjected to phenotypic characterization (antimicrobial susceptibility, growth, sporulation ability and toxin production) and WGS analysis to evaluate clonality and modifications associated with resistance., Results: Resistance to fidaxomicin arose in isolates from the recurrences of CDI (MIC: 16 mg/L). WGS analysis showed a close genetic link between strains suggestive of relapses in this patient. A T3428G mutation in the rpoB gene might be associated with fidaxomicin resistance. The resistance was associated with defects in growth, sporulation and production of toxins. A review of the literature found only three previous fidaxomicin-resistant C. difficile clinical strains., Conclusions: Although rarely reported, resistance to fidaxomicin may quickly emerge in vivo after a single course of treatment. This observation supports the need for prospective surveillance of the susceptibility of C. difficile to treatment antibiotics. However, the clinical relevance of fidaxomicin resistance still needs to be elucidated, particularly due to its apparent rareness and associated fitness cost., (© The Author(s) 2023. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Introduction: Clostridioides difficile infection (CDI) is a leading cause of healthcare-associated (HA) diarrhoea. We retrospectively investigated data from a comprehensive, multidisciplinary C. difficile surveillance programme focusing on hospitalized patients in a tertiary Irish hospital over 10 years., Methods: Data from 2012 to 2021 were extracted from a centralized database, including patient demographics, admission, case and outbreak details, ribotypes (RTs), and (since 2016) antimicrobial exposures and CDI treatments. Counts of CDI by origin of infection were explored using ꭓ 2 analyses, Poisson regression was used to investigate trends in rates of CDI and possible risk factors. Time to recurrent CDI was examined by a Cox proportional hazards regression., Results: Over 10 years, 954 CDI patients had a 9% recurrent CDI rate. CDI testing requests occurred in only 22% of patients. Most CDIs were HA (82.2%) and affected females (odds ratio: 2.3, P<0.01). Fidaxomicin significantly reduced the hazard ratio of time to recurrent CDI. No trends in HA-CDI incidence were observed despite key time-point events and increasing hospital activity. In 2021, community-associated (CA)-CDI increased. RTs did not differ for HA versus CA for the most common RTs (014, 078, 005 and 015). Average length-of-stay differed significantly between HA (67.1 days) and CA (14.6 days) CDI., Conclusion: HA-CDI rates remained unchanged despite key events and increased hospital activity, whereas by 2021, CA-CDI was at its highest in a decade. The convergence of CA and HA RTs, and the proportion of CA-CDI, question the relevance of current case definitions when increasingly patients receive hospital care without an overnight hospital stay., (Copyright © 2023 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Background: Awareness and compliance with international guidelines for diagnosis and clinical management of Clostridioides difficile infection (CDI) are unknown., Aim: To compare the awareness and compliance with the recommended strategies for diagnosis and clinical management of CDI across Europe in 2018-2019., Methods: Hospital sites and their associated community practices across 12 European countries completed an online survey in 2018-2019, to report on their practices in terms of surveillance, prevention, diagnosis, and treatment of CDI. Responses were collected from 105 hospitals and 39 community general practitioners (GPs)., Findings: Hospital sites of 11 countries reported participation in national surveillance schemes compared with six countries for international schemes. The European Society of Clinical Microbiology and Infectious Diseases (ESCMID)-recommended CDI testing methodologies were used by 82% (86/105) of hospitals, however countries reporting the highest incidence of CDI used non-recommended tests. Over 75% (80/105) of hospitals were aware of the most recent European CDI treatment guidelines at the time of this survey compared with only 26% (10/39) of surveyed GPs. However, up to 15% (16/105) of hospitals reported using the non-recommended metronidazole for recurrent CDI cases, sites in countries with lower awareness of CDI treatment guidelines. Only 37% (39/105) of hospitals adopted contact isolation precautions in case of suspected CDI., Conclusion: Good awareness of guidelines for the management of CDI was observed across the surveyed European hospital sites. However, low compliance with diagnostic testing guidelines, infection control measures for suspected CDI, and insufficient awareness of treatment guidelines continued to be reported in some countries., (Copyright © 2022 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Recently, the recommendations for the treatment of Clostridioides difficile infection (CDI) have been updated. However, in addition to the clinical efficacy data, the drug of choice should ideally represent optimal antimicrobial stewardship, with an emphasis on rapid restoration of the gut microbiota to minimize the risk of infection relapses. Oral administration of metronidazole results in low concentration in stool, and interaction with fecal microbiota reduces its antimicrobial bioactivity. Reported elevated minimum inhibitory concentrations of metronidazole in epidemic C. difficile ribotypes and the emergence of plasmid-mediated resistance to metronidazole represent additional potential risks for clinical failure. If metronidazole is the only CDI treatment option, antimicrobial susceptibility testing on agar containing heme should be performed in C. difficile isolate. Compared with metronidazole, oral vancomycin and fidaxomicin reach very high concentrations in the stool, and therefore can quickly reduce C. difficile shedding. Health care facilities with higher CDI incidence and/or occurrence of epidemic ribotypes should not use metronidazole because prolonged C. difficile shedding can increase the risk for further C. difficile transmission. Only fidaxomicin has a narrow spectrum of antimicrobial activity, which might be, together with persistence on spores, the main contributing factor to reduce the recurrent CDI rates., Competing Interests: Declaration of Competing Interest M.K. and E.J.K. have no competing interests to declare. M.H.W. received lecture fees from Merck, Pfizer, and Seres and consultation fees from AiCuris, Bayer, Crestone, Da Volterra, Deinove, EnteroBiotix, The European Tissue Symposium, Ferring, GSK, Menarini, Merck, Nestlé, Paratek, Pfizer, Phico Therapeutics, Qpex Biopharma, Seres, Surface Skins, Summit, and Vaxxilon/Idorsia., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

BackgroundWhile human-to-human transmission of Clostridioides difficile occurs often, other infection sources, including food, animals and environment, are under investigation.AimWe present a large study on C. difficile in a food item in Europe, encompassing 12 European countries (Austria, France, Greece, Ireland, Italy, the Netherlands, Poland, Slovakia, Spain, Sweden, Romania and the United Kingdom).MethodsPotato was selected because of availability, ease of sampling and high C. difficile positivity rates. Identical protocols for sampling and isolation were used, enabling a direct comparison of the C. difficile positivity rate.ResultsFrom C. difficile -positive potato samples (33/147; 22.4%), we obtained 504 isolates, grouped into 38 PCR ribotypes. Positivity rates per country varied (0-100%) and were at least 10% in 9/12 countries. No geographical clustering of samples with high positivity rates or in PCR ribotype distribution was observed. The most frequently detected PCR ribotypes (014/020, 078/126, 010 and 023) are also commonly reported in Europe among human clinically relevant isolates, in animal isolates and in the environment. Whole genome sequencing revealed several genetically related strain pairs (Spain/RT126, France/RT010, Austria and Sweden/RT276) and a cluster of very similar strains in RT078/126.ConclusionOur results suggest, the high potato contamination rates could have public health relevance. They indicate potatoes can serve as a vector for introducing C. difficile spores in the household environment, where the bacterium can then multiply in sensitive hosts with disrupted or unmature microbiota. Potato contamination with PCR ribotypes shared between humans, animals and soil is supportive of this view.

Background: There are inconsistent data on the risk factors for Clostridium difficile infection (CDI) in the literature. Aims: To use two C. difficile infection (CDI) case-control study groups to compare risk factors in hospitalized patients with diarrhea across different countries. Methods: A multi-center group of CDI cases/controls were identified by standardized testing from seven countries from the prior EUropean, multi-center, prospective bi-annual point prevalence study of CLostridium difficile Infection in hospitalized patients with Diarrhea (EUCLID). A second group of CDI cases/controls was identified from a single center in Germany [parallel study site (PSS)]. Data were extracted from the medical notes to assess CDI risk factors. Univariate analyses and multivariate logistic regression models were used to identify and compare risk factors between the two groups. Results: There were 253 and 158 cases and 921 and 584 controls in the PSS and EUCLID groups, respectively. Significant variables from univariate analyses in both groups were age ≥65, number of antibiotics (OR 1.2 for each additional antibiotic) and prior hospital admission (all p < 0.001). Congestive heart failure, diabetes, admission from assisted living or Emergency Department, proton pump inhibitors, and chronic renal disease were significant in PSS (all p < 0.05) but not EUCLID. Dementia and admitted with other bacterial diseases were significant in EUCLID ( p < 0.05) but not PSS. Following multivariate analyses, age ≥ 65, number of antibiotics and prior hospital admission were consistently identified as CDI risk factors in each individual group and combined datasets. Conclusion: Our results show that the same CDI risk factors were identified across datasets. These were age ≥ 65 years, antibiotic use and prior hospital admission. Importantly, the odds of developing CDI increases with each extra antibiotic prescribed., (Copyright © 2020 Davies, Lawrence, Berry, Davis, Yu, Cai, Gonzalez, Prantner, Kurcz, Macovei, Pituch, Nováková, Nyč, Gärtner, Berger, Oleastro, Cornely, Vehreschild, Pedneault and Wilcox.)

Background: Clostridioides difficile infection (CDI) remains a high burden worldwide. DAV131A, a novel adsorbent, reduces residual gut antimicrobial levels, reducing CDI risk in animal models., Objectives: We used a validated human gut model to investigate the efficacy of DAV131A in preventing moxifloxacin-induced CDI., Methods: C. difficile (CD) spores were inoculated into two models populated with pooled human faeces. Moxifloxacin was instilled (43 mg/L, once daily, 7 days) alongside DAV131A (5 g in 18 mL PBS, three times daily, 14 days, Model A), or PBS (18 mL, three times daily, 14 days, Model B). Selected gut microbiota populations, CD total counts, spore counts, cytotoxin titre and antimicrobial concentrations (HPLC) were monitored daily. We monitored for reduced susceptibility of CD to moxifloxacin. Growth of CD in faecal filtrate and medium in the presence/absence of DAV131A, or in medium pre-treated with DAV131A, was also investigated., Results: DAV131A instillation reduced active moxifloxacin levels to below the limit of detection (50 ng/mL), and prevented microbiota disruption, excepting Bacteroides fragilis group populations, which declined by ∼3 log10 cfu/mL. DAV131A delayed onset of simulated CDI by ∼2 weeks, but did not prevent CD germination and toxin production. DAV131A prevented emergence of reduced susceptibility of CD to moxifloxacin. In batch culture, DAV131A had minor effects on CD vegetative growth, but significantly reduced toxin/spores (P < 0.005)., Conclusions: DAV131A reduced moxifloxacin-induced microbiota disruption and emergence of antibiotic-resistant CD. Delayed onset of CD germination and toxin production indicates further investigations are warranted to understand the clinical benefits of DAV131A in CDI prevention., (© The Author(s) 2020. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Clostridium difficile infection (CDI) has been primarily treated with metronidazole or vancomycin. High recurrence rates, the emergence of epidemic PCR ribotypes (RTs) and the introduction of fidaxomicin in Europe in 2011 necessitate surveillance of antimicrobial resistance and CDI epidemiology. The ClosER study monitored antimicrobial susceptibility and geographical distribution of C. difficile RTs pre- and post-fidaxomicin introduction. From 2011 to 2016, 28 European countries submitted isolates or faecal samples for determination of PCR ribotype, toxin status and minimal inhibitory concentrations (MICs) of metronidazole, vancomycin, rifampicin, fidaxomicin, moxifloxacin, clindamycin, imipenem, chloramphenicol and tigecycline. RT diversity scores for each country were calculated and mean MIC results used to generate cumulative resistant scores (CRSs) for each isolate and country. From 40 sites, 3499 isolates were analysed, of which 95% (3338/3499) were toxin positive. The most common of the 264 RTs isolated was RT027 (mean prevalence 11.4%); however, RT prevalence varied greatly between countries and between years. The fidaxomicin geometric mean MIC for years 1-5 was 0.04 mg/L; only one fidaxomicin-resistant isolate (RT344) was submitted (MIC ≥ 4 mg/L). Metronidazole and vancomycin geometric mean MICs were 0.46 mg/L and 0.70 mg/L, respectively. Of prevalent RTs, RT027, RT017 and RT012 demonstrated resistance or reduced susceptibility to multiple antimicrobials. RT diversity was inversely correlated with mean CRS for individual countries (Pearson coefficient r = - 0.57). Overall, C. difficile RT prevalence remained stable in 2011-2016. Fidaxomicin susceptibility, including in RT027, was maintained post-introduction. Reduced ribotype diversity in individual countries was associated with increased antimicrobial resistance.

Scope: Clostridium difficile infection (CDI) is the most important infective cause of healthcare-associated diarrhoea in high income countries and one of the most important healthcare-associated pathogens in both Europe and the United States. It is associated with high morbidity and mortality resulting in both societal and financial burden. A significant proportion of this burden is potentially preventable by a combination of targeted infection prevention and control measures and antimicrobial stewardship. The aim of this guidance document is to provide an update on recommendations for prevention of CDI in acute care settings to provide guidance to those responsible for institutional infection prevention and control programmes., Methods: An expert group was set up by the European society of clinical microbiology and infectious diseases (ESCMID) Study Group for C. difficile (ESGCD), which performed a systematic review of the literature on prevention of CDI in adults hospitalized in acute care settings and derived respective recommendations according to the GRADE approach. Recommendations are stratified for both outbreak and endemic settings., Questions Addressed by the Guideline and Recommendations: This guidance document provides thirty-six statements on strategies to prevent CDI in acute care settings, including 18 strong recommendations. No recommendation was provided for three questions., (Copyright © 2018 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Guidelines recommend the use of an algorithm for the laboratory diagnosis of Clostridium difficile infection (CDI). Enzyme immunoassays (EIAs) detecting C. difficile toxins cannot be used as standalone tests due to suboptimal sensitivity, and molecular tests suffer from nonspecificity by detecting colonization. Sensitive immunoassays have recently been developed to improve and simplify CDI diagnosis. Assays detecting CD toxins have been developed using single-molecule array (SIMOA) technology. SIMOA performance was assessed relative to a laboratory case definition of CDI defined by positive glutamate dehydrogenase (GDH) screen and cell cytotoxicity neutralizing assay (CCNA). Samples were tested with SIMOA assays and a commercial toxin EIA to compare performance, with discrepancy resolution using a commercial nucleic acid-based test and a second cell cytotoxicity assay. The SIMOA toxin A and toxin B assays showed limits of detection of 0.6 and 2.9 pg/ml, respectively, and intra-assay coefficients of variation of less than 10%. The optimal clinical thresholds for the toxin A and toxin B assays were determined to be 22.1 and 18.8 pg/ml, respectively, with resultant sensitivities of 84.8 and 95.5%. In contrast, a high-performing EIA toxin test had a sensitivity of 71.2%. Thus, the SIMOA assays detected toxins in 24% more samples with laboratory-defined CDI than the high performing toxin EIA (95% [63/66] versus 71% [47/66]). This study shows that SIMOA C. difficile toxin assays have a higher sensitivity than currently available toxin EIA and have the potential to improve CDI diagnosis., (Copyright © 2018 Banz et al.)

Objectives: Until the introduction of fidaxomicin, antimicrobial treatment for Clostridium difficile infection (CDI) was limited to metronidazole and vancomycin. The changing epidemiology of CDI and the emergence of epidemic C. difficile PCR ribotype 027 necessitate continued surveillance to identify shifts in antibiotic susceptibility. ClosER, currently the largest pan-European epidemiological study of C. difficile ribotype distribution and antibiotic susceptibility, aimed to undertake antimicrobial resistance surveillance pre- and post-introduction of fidaxomicin., Methods: Between July 2011 and July 2014, 39 sites across 22 European countries submitted 2830 C. difficile isolates for ribotyping, toxin testing and susceptibility testing to metronidazole, vancomycin, fidaxomicin, rifampicin, moxifloxacin, clindamycin, imipenem, chloramphenicol and tigecycline., Results: Ribotypes 027, 014, 001, 078, 020, 002, 126, 015 and 005 were most frequently isolated, and emergent ribotypes 198 and 356 were identified in Hungary and Italy, respectively. All isolates were susceptible to fidaxomicin, with scarce resistance to metronidazole (0.2%, 6/2694), vancomycin (0.1%, 2/2694) and tigecycline (0%). Rifampicin, moxifloxacin and clindamycin resistance was evident in multiple ribotypes. Lack of ribotype diversity correlated with greater antimicrobial resistance. Epidemic ribotypes (027/001) were associated with multiple antimicrobial resistance, and ribotypes 017, 018 and 356 with high-level resistance. Additional factors may also influence local ribotype prevalence., Conclusions: Fidaxomicin susceptibility was retained post-introduction, and resistance to metronidazole and vancomycin was rare. Continued surveillance is needed, with more accurate classification and clarification of ribotype subtypes to further understand their role in the spread of resistance. Other factors may also influence changes in prevalence of C. difficile ribotypes with reduced antibiotic susceptibility., (Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Background: The increasing incidence of Clostridium difficile infections (CDI) in healthcare settings in Europe since 2003 has affected both patients and healthcare systems. The implementation of effective CDI surveillance is key to enable monitoring of the occurrence and spread of C. difficile in healthcare and the timely detection of outbreaks., Aims: The aim of this review is to provide a summary of key components of effective CDI surveillance and to provide some practical recommendations. We also summarize the recent and current national CDI surveillance activities, to illustrate strengths and weaknesses of CDI surveillance in Europe., Sources: For the definition of key components of CDI surveillance, we consulted the current European Society of Clinical Microbiology and Infectious Diseases (ESCMID) CDI-related guidance documents and the European Centre for Disease Prevention and Control (ECDC) protocol for CDI surveillance in acute care hospitals. To summarize the recent and current national CDI surveillance activities, we discussed international multicentre CDI surveillance studies performed in 2005-13. In 2017, we also performed a new survey of existing CDI surveillance systems in 33 European countries., Content: Key components for CDI surveillance are appropriate case definitions of CDI, standardized CDI diagnostics, agreement on CDI case origin definition, and the presentation of CDI rates with well-defined numerators and denominators. Incorporation of microbiological data is required to provide information on prevailing PCR ribotypes and antimicrobial susceptibility to first-line CDI treatment drugs. In 2017, 20 European countries had a national CDI surveillance system and 21 countries participated in ECDC-coordinated CDI surveillance. Since 2014, the number of centres with capacity for C. difficile typing has increased to 35 reference or central laboratories in 26 European countries., Implications: Incidence rates of CDI, obtained from a standardized CDI surveillance system, can be used as an important quality indicator of healthcare at hospital as well as country level., (Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.)

Some strains of Clostridium difficile produce a binary toxin, in addition to the main C. difficile virulence factors (toxins A and B). There have been conflicting reports regarding the role of binary toxin and its relationship to the severity of C. difficile infection (CDI). Samples, isolates and clinical data were collected as part of a prospective multicentre diagnostic study. Clostridium difficile isolates (n = 1259) were tested by polymerase chain reaction (PCR) assay to detect binary toxin genes cdtA and cdtB. The PCR binary toxin gene results were compared with clinical severity and outcome data, including 30-day all-cause mortality. The 1259 isolates corresponded to 1083 different patients (October 2010 to September 2011). The prevalence of binary toxin positive strains was significantly higher in faecal samples with detectable toxin A/B than in those without toxin but that were positive by cytotoxigenic culture (26.3% vs. 10.3%, p < 0.001). The presence of binary toxin correlated moderately with markers of CDI severity (white cell count, serum albumin concentration and serum creatinine concentration). However, the risk ratio for all-cause mortality was 1.68 for binary toxin positive patients and patients were significantly less likely to survive if they had CDI caused by a binary toxin gene positive strain, even after adjusting for age (p < 0.001). The presence of binary toxin genes does not predict the clinical severity of CDI, but it is significantly associated with the risk of all-cause mortality.

Purpose of Review: Clostridium difficile infection has attained high prominence given its prevalence and impacts on patients and healthcare institutions. Multiple new approaches to the prevention and treatment of C. difficile infection (CDI) are undergoing clinical trials., Recent Findings: Bezlotoxumab is a monoclonal antibody against toxin B that has successfully completed phase III studies, demonstrating a significant reduction in recurrent CDI when given with standard of care antibiotics. Antibiotics under development include cadazolid and ridinilazole, whereas surotomycin has had disappointing phase III results. Multiple live biotherapeutics are being developed, including freeze thawed and encapsulated versions of faecal microbiota transplantation to improve the practicality of treating patients with recurrent CDI. Alternatives to faecal microbiota transplantation, that aim to improve safety, including a microbial suspension, RBX2660, and a complex spore formulation, SER-109, have progressed to phase II studies. A nontoxigenic C. difficile strain has also shown promise to prevent recurrent CDI. In addition, three C. difficile vaccines have progressed to phase II/III clinical trials., Summary: The diverse approaches to treating and preventing CDI offer substantial promise that new treatment options will soon emerge, particular ones that reduce the risk of recurrences.

Lack of standardised Clostridium difficile testing is a potential confounder when comparing infection rates. We used an observational, systematic, prospective large-scale sampling approach to investigate variability in C. difficile sampling to understand C. difficile infection (CDI) incidence rates. In-patient and institutional data were gathered from 60 European hospitals (across three countries). Testing methodology, testing/CDI rates and case profiles were compared between countries and institution types. The mean annual CDI rate per hospital was lowest in the UK and highest in Italy (1.5 vs. 4.7 cases/10,000 patient bed days [pbds], p < 0.001). The testing rate was highest in the UK compared with Italy and France (50.7/10,000 pbds vs. 31.5 and 30.3, respectively, p < 0.001). Only 58.4 % of diarrhoeal samples were tested for CDI across all countries. Overall, only 64 % of hospitals used recommended testing algorithms for laboratory testing. Small hospitals were significantly more likely to use standalone toxin tests (SATTs). There was an inverse correlation between hospital size and CDI testing rate. Hospitals using SATT or assays not detecting toxin reported significantly higher CDI rates than those using recommended methods, despite testing similar testing frequencies. These data are consistent with higher false-positive rates in such (non-recommended) testing scenarios. Cases in Italy and those diagnosed by SATT or methods NOT detecting toxin were significantly older. Testing occurred significantly earlier in the UK. Assessment of testing practice is paramount to the accurate interpretation and comparison of CDI rates., Competing Interests: Compliance with ethical standards Funding This study was initiated and funded by Sanofi Pasteur (France). Conflict of interest KD has received honoraria from Astellas Pharma Europe and grant support from Astellas Pharma Europe, bioMérieux, Pfizer and Sanofi Pasteur. GD has received grant support from Astellas Pharma Europe, bioMérieux, Pfizer and Sanofi Pasteur. FB is an advisory board member for and/or has received scientific grants from Merck, Cubist, Sanofi Pasteur, Da Volterra, Pfizer and Astellas. FB has received scientific grants from bioMérieux, Alere, Cubist, Astellas, Orion Diagnostica, Greiner and Mobidiag. CE has received conference funding from Alere and Astellas and honoraria from Astellas. NP has received honoraria as a speaker for MSD, Pfizer, Astellas, Novartis and Angelini, and received honoraria as a member of the scientific advisory board for MSD, Pfizer, Astellas, The Medicines Company, Zambon, Carefusion and Achaogen. MHW has received: consulting fees from Abbott Laboratories, Actelion, Astellas, Astra-Zeneca, Bayer, bioMérieux, Cerexa, Cubist, Durata, The European Tissue Symposium, The Medicines Company, MedImmune, Merck, Motif Biosciences, Nabriva, Optimer, Paratek, Pfizer, Qiagen, Roche, Sanofi Pasteur, Seres, Summit and Synthetic Biologics; lecture fees from Abbott, Alere, Astellas, Astra-Zeneca, Merck, Pfizer and Roche; and grant support from Abbott, Actelion, Astellas, bioMérieux, Cubist, Da Volterra, Sanofi Pasteur, Seres, Summit, The European Tissue Symposium and Merck. Ethical approval This surveillance study was granted ethical approval by the University of Leeds (SoMREC13032) for UK data collection and European-wide analysis, and by the National Institute for Infectious Diseases ‘Spallanzani’, Rome for Italian data collection. Ethical approval was not required in France. Informed consent Informed consent is not required for this surveillance study, in line with the ethical approvals shown above.

Clostridium difficile is a leading nosocomial pathogen and molecular typing is a crucial part of monitoring its occurrence and spread. Over a three-year period (2013-2015), clinical C. difficile isolates from 32 Czech hospitals were collected for molecular characterisation. Of 2201 C. difficile isolates, 177 (8%) were non-toxigenic, 2024 (92%) were toxigenic (tcdA and tcdB) and of these, 677 (33.5%) carried genes for binary toxin production (cdtA, cdtB). Capillary-electrophoresis (CE) ribotyping of the 2201 isolates yielded 166 different CE-ribotyping profiles, of which 53 were represented by at least two isolates for each profile. Of these, 29 CE-ribotyping patterns were common to the Leeds-Leiden C. difficile reference strain library and the WEBRIBO database (83.7% isolates), and 24 patterns were recognized only by the WEBRIBO database (11.2% isolates). Isolates belonging to these 53 CE-ribotyping profiles comprised 94.9% of all isolates. The ten most frequent CE-ribotyping profiles were 176 (n=588, 26.7%), 001 (n=456, 20.7%), 014 (n=176, 8%), 012 (n=127, 5.8%), 017 (n=85, 3.9%), 020 (n=68, 3.1%), 596 (n=55, 2.5%), 002-like (n=45, 2.1%), 010 (n=35, 1.6%) and 078 (n=34, 1.6%). Multi-locus sequence typing (MLST) of seven housekeeping genes performed in one isolate of each of 53 different CE-ribotyping profiles revealed 40 different sequence types (STs). We conclude that molecular characterisation of Czech C. difficile isolates revealed a high diversity of CE-ribotyping profiles; the prevailing RTs were 001 (20.7%) and 176 (027-like, 26.7%)., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

In 2009 the first European Society of Clinical Microbiology and Infectious Diseases (ESCMID) guideline for diagnosing Clostridium difficile infection (CDI) was launched. Since then newer tests for diagnosing CDI have become available, especially nucleic acid amplification tests. The main objectives of this update of the guidance document are to summarize the currently available evidence concerning laboratory diagnosis of CDI and to formulate and revise recommendations to optimize CDI testing. This update is essential to improve the diagnosis of CDI and to improve uniformity in CDI diagnosis for surveillance purposes among Europe. An electronic search for literature concerning the laboratory diagnosis of CDI was performed. Studies evaluating a commercial laboratory test compared to a reference test were also included in a meta-analysis. The commercial tests that were evaluated included enzyme immunoassays (EIAs) detecting glutamate dehydrogenase, EIAs detecting toxins A and B and nucleic acid amplification tests. Recommendations were formulated by an executive committee, and the strength of recommendations and quality of evidence were graded using the Grades of Recommendation Assessment, Development and Evaluation (GRADE) system. No single commercial test can be used as a stand-alone test for diagnosing CDI as a result of inadequate positive predictive values at low CDI prevalence. Therefore, the use of a two-step algorithm is recommended. Samples without free toxin detected by toxins A and B EIA but with positive glutamate dehydrogenase EIA, nucleic acid amplification test or toxigenic culture results need clinical evaluation to discern CDI from asymptomatic carriage., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: Clostridium difficile infection (CDI) is a global healthcare problem. Recent evidence suggests that the availability of iron may be important for C. difficile growth. This study evaluated the comparative effects of iron-depleted (1% Fe(3+) saturated) bovine apo-lactoferrin (apo-bLf) and iron-saturated (85% Fe(3+) saturated) bovine holo-lactoferrin (holo-bLf) in a human in vitro gut model that simulates CDI., Methods: Two parallel triple-stage chemostat gut models were inoculated with pooled human faeces and spiked with C. difficile spores (strain 027 210, PCR ribotype 027). Holo- or apo-bLf was instilled (5 mg/mL, once daily) for 35 days. After 7 days, clindamycin was instilled (33.9 mg/L, four times daily) to induce simulated CDI. Indigenous microflora populations, C. difficile total counts and spores, cytotoxin titres, short chain fatty acid concentrations, biometal concentrations, lactoferrin concentration and iron content of lactoferrin were monitored daily., Results: In the apo-bLf model, germination of C. difficile spores occurred 6 days post instillation of clindamycin, followed by rapid vegetative cell proliferation and detectable toxin production. By contrast, in the holo-bLf model, only a modest vegetative cell population was observed until 16 days post antibiotic administration. Notably, no toxin was detected in this model. In separate batch culture experiments, holo-bLf prevented C. difficile vegetative cell growth and toxin production, whereas apo-bLf and iron alone did not., Conclusions: Holo-bLf, but not apo-bLf, delayed C. difficile growth and prevented toxin production in a human gut model of CDI. This inhibitory effect may be iron independent. These observations suggest that bLf in its iron-saturated state could be used as a novel preventative or treatment strategy for CDI., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)

Objectives: The aim of this study was to evaluate the susceptibilities of Clostridium difficile isolates to cadazolid, a novel antibiotic for the treatment of C. difficile infection., Methods: Ribotyping and susceptibilities were determined for C. difficile isolates from a multicentre, double-blind, Phase 2 study of oral cadazolid in patients with C. difficile infection (NCT01222702, ClinicalTrials.gov; EudraCT 2010-020941-29, European Clinical Trials Database). Patients were randomized to receive 250, 500 or 1000 mg of cadazolid twice daily or 125 mg of vancomycin four times daily, for 10 days. MICs of cadazolid, vancomycin, fidaxomicin, linezolid and moxifloxacin were determined at baseline for all patients and post-baseline for patients with clinical failure or recurrence, using the agar dilution method., Results: Seventy-eight of 84 patients had an evaluable toxigenic C. difficile isolate at baseline. The most frequent PCR ribotype was 027 (15.4%). Cadazolid MICs for baseline isolates (including epidemic strain 027) ranged from 0.06 to 0.25 mg/L. Baseline cadazolid MICs were similar to those of fidaxomicin and lower than those of vancomycin, linezolid and moxifloxacin. For each clinical outcome group (clinical cure, clinical failure, sustained clinical response and clinical failure or recurrence), the baseline cadazolid MIC range was 0.06-0.25 mg/L. Mean (min-max) cadazolid faecal concentration (μg/g) on day 5 was 884 (101-2710), 1706 (204-4230) and 3226 (1481-12 600) for the doses 250, 500 and 1000 mg, respectively., Conclusions: For all cadazolid doses, the faecal concentration was in excess of several thousand-fold the MIC90 for C. difficile. The MIC of cadazolid for all C. difficile isolates, including epidemic strains, was low and in the same narrow range regardless of treatment outcome., (© The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)

Cadazolid, a novel fluoroquinolone-oxazolidinone antibiotic, exhibits potent in vitro activity against Clostridium difficile, including the epidemic BI/NAP1/027 strain. This multicenter, randomized, double-blind, active reference group, phase 2 study evaluated the efficacy and safety of oral cadazolid in treatment of adult patients with C. difficile infection (CDI). Eligible patients with first occurrence/first recurrence of CDI were randomized 1:1:1:1 to 250, 500, or 1,000 mg cadazolid twice daily (BID) or oral 125 mg vancomycin four times daily (QID) for 10 days. The primary endpoint was clinical cure at test of cure (48 ± 24 h after the end of treatment; modified intent-to-treat population), defined as resolution of diarrhea with no further CDI treatment required. Secondary endpoints included recurrence rate, sustained clinical response (clinical cure without recurrence), and time to diarrhea resolution. Of 84 patients enrolled, 20, 22, 20, and 22 received 250, 500, or 1,000 mg cadazolid BID or 125 mg vancomycin QID, respectively. The primary endpoint was achieved in 76.5% (80% confidence interval [CI], 58.4, 89.3), 80.0% (63.9, 91.0), 68.4% (51.1, 82.5), and 68.2% (52.3, 81.3) of patients, respectively. There was no evidence of a cadazolid dosage-dependent response. Each dosage of cadazolid resulted in a lower recurrence rate than with vancomycin (18.2 to 25.0% versus 50%). Consequently, higher sustained clinical response rates were observed with cadazolid (46.7 to 60.0%) than with vancomycin (33.3%). The times to diarrhea resolution were similar for cadazolid and vancomycin. Cadazolid was well tolerated, with no safety signal observed. The results of this phase 2 study support further clinical development of cadazolid. (This study has been registered in the United States at ClinicalTrials.gov under registration no. NCT01222702 and in Europe with the European Medicines Agency under registration no. EUDRA-CT 2010-020941-29.)., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Background: Fidaxomicin treatment reduces the risk of recurrent Clostridium difficile infection (CDI) compared with vancomycin. Extending duration of fidaxomicin therapy may further reduce recurrence. We compared the efficacy of four extended fidaxomicin regimens in an in vitro model of CDI., Methods: Four gut models were primed with human faeces, spiked with C. difficile spores (PCR ribotype 027) and clindamycin instilled (33.9 mg/L, four-times daily, 7 days) to induce simulated CDI. Four extended fidaxomicin treatment regimens were evaluated: model 1, 20 days, 200 mg/L twice daily; model 2, 5 days 200 mg/L twice daily, 5 days rest, 5 days 200 mg/L twice daily; model 3, 5 days 200 mg/L twice daily, 5 days rest, 10 days 200 mg/L once daily; and model 4, 5 days 200 mg/L twice daily, 20 days 200 mg/L once every other day. C. difficile populations, toxin, gut microbiota and antimicrobial levels were monitored daily., Results: All fidaxomicin regimens successfully resolved simulated CDI without recurrence. Five days of fidaxomicin instillation was barely sufficient to resolve CDI (models 2-4). A second pulse or tapered dosing further reduced C. difficile and toxin detection. All regimens were sparing of microbiota, affecting only enterococci and bifidobacteria. Pulsed or tapered regimens allowed greater bifidobacteria recovery than the extended (20 day) regimen. Bioactive fidaxomicin persisted throughout the experiment in all models at concentrations inhibitory to C. difficile., Conclusions: Pulsed or tapered fidaxomicin regimens may enhance suppression of C. difficile whilst allowing microbiota recovery; clinical studies are required to ascertain the potential of this approach in further reducing recurrent CDI., (© The Author 2015. 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.)

Background: Clostridium difficile infection (CDI) is a leading cause of diarrhoea in health care settings with symptoms ranging from mild and self-limiting to life threatening. SMT19969 is a novel, non-absorbable antibiotic currently under development for the treatment of CDI. Here we report the results from a Phase I study., Methods: A double-blind, randomized, placebo-controlled study assessing safety and tolerability of single and multiple oral doses of SMT19969 in healthy volunteers. Pharmacokinetic assessments included blood and faecal sampling. The effect of food on systemic exposure and analysis of the gut microbiota were also included., Results: Fifty-six healthy male subjects were enrolled. Following single oral doses of up to 2,000 mg in the fasted state, plasma concentrations of SMT19969 were generally below the lower limit of quantification. In the fed state levels ranged from 0.102 to 0.296 ng/mL after single dosing and after repeat dosing at Day 10 from 0.105 to 0.305 ng/mL. Following single and multiple oral doses of SMT19969, mean daily faecal concentrations increased with increasing dose level and were significantly above the typical MIC range for C. difficile (0.06-0.5 μg/mL). At 200 mg BID, mean (± SD) faecal concentrations of 1,466 (±547) μg/g and 1,364 (±446) μg/g were determined on days 5 and 10 of dosing respectively. No notable metabolites were detected in faeces. Overall, all doses of SMT19969 were well tolerated both as single oral doses or BID oral doses for 10 days. The majority (88%) of adverse events (AEs) were classified as gastrointestinal disorders and were mild in severity, resolving without treatment. The gut microbiota was analysed in the multiple dose groups with minimal changes observed in the bacterial groups analysed except for total clostridia which were reduced to below the limit of detection by day 4 of dosing., Conclusions: Oral administration of SMT19969 was considered safe and well tolerated and was associated with negligible plasma concentrations after single and multiple doses. In addition, minimal disruption of normal gut microbiota was noted, confirming the highly selective spectrum of the compound. These results support the further clinical development of SMT19969 as an oral therapy for CDI., Trial Registration: Current Controlled Trials. ISRCTN10858225 .

Objectives: We investigated the efficacy of the cyclic lipopeptide surotomycin in treating clindamycin-induced Clostridium difficile infection (CDI) using an in vitro gut model., Methods: Two three-stage chemostat gut models were inoculated with human faeces, spiked with C. difficile spores (∼10(7) cfu/mL, PCR ribotype 027 or 001). Clindamycin (33.9 mg/L, four times daily for 7 days) was dosed to induce CDI. Following high-level toxin production, surotomycin (250 mg/L, twice daily for 7 days) was instilled. Microflora populations, C. difficile vegetative cells and spores, cytotoxin titres and antimicrobial levels (LC-MS/MS and bioassay) were determined. The emergence of C. difficile and enterococci with reduced susceptibility to surotomycin was monitored on breakpoint agar (4 × MIC)., Results: Counts of viable C. difficile were reduced to near the limit of detection on Days 1 and 3 of surotomycin instillation, and cytotoxin was undetectable on Days 3 and 4 of surotomycin instillation in the 027 and 001 models, respectively. Recurrence of vegetative growth and toxin production occurred 11 days (001 model) and 15 days (027 model) after surotomycin instillation had ceased, and remained for the duration of the experiment. Surotomycin instillation decreased populations of bifidobacteria, clostridia, enterococci and lactobacilli, but was sparing of Bacteroides fragilis group populations. All enumerated organisms had recovered to steady-state levels by 3 weeks post-surotomycin instillation. No evidence of the emergence of reduced susceptibility to surotomycin was observed., Conclusions: Surotomycin successfully reduced C. difficile vegetative cell counts and toxin levels in the gut model and was sparing of B. fragilis group populations. There was no evidence of decreased susceptibility to surotomycin during exposure or post-exposure., (© 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.)

Background: Clostridium difficile infection (CDI) remains an infection control challenge, especially when environmental spore contamination and suboptimal cleaning may increase transmission risk., Aim: To substantiate the long-term effectiveness throughout a stroke rehabilitation unit (SRU) of deep cleaning and hydrogen peroxide decontamination (HPD), following a high incidence of CDI., Methods: Extensive environmental sampling (342 sites on each occasion) for C. difficile using sponge wipes was performed: before and after deep cleaning with detergent/chlorine agent; immediately following HPD; and on two further occasions, 19 days and 20 weeks following HPD. C. difficile isolates underwent polymerase chain reaction ribotyping and multi-locus variable repeat analysis (MLVA)., Findings: C. difficile was recovered from 10.8%, 6.1%, 0.9%, 0% and 3.5% of sites at baseline, following deep cleaning, immediately after HPD, and 19 days and 20 weeks after HPD, respectively. C. difficile ribotypes recovered after deep cleaning matched those from CDI cases in the SRU during the previous 10 months. Similarly, 10/12 of the positive sites identified at 20 weeks post-HPD harboured the same C. difficile ribotype (002) and MLVA pattern as the isolate from the first post-HPD CDI case. CDI incidence [number of cases on SRU per 10 months (January-October 2011)] declined from 20 before to seven after the intervention., Conclusion: HPD, after deep cleaning with a detergent/chlorine agent, was highly effective for removing environmental C. difficile contamination. Long-term follow-up demonstrated that a CDI symptomatic patient can rapidly recontaminate the immediate environment. Determining a role for HPD should include long-term cost-effectiveness evaluations., (Copyright © 2014 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Objectives: We investigated the in vitro activity of cadazolid against 100 Clostridium difficile isolates and its efficacy in a simulated human gut model of C. difficile infection (CDI)., Methods: MICs of cadazolid, metronidazole, vancomycin, moxifloxacin and linezolid were determined using agar incorporation for 100 C. difficile isolates, including 30 epidemic strains (ribotypes 027, 106 and 001) with reduced metronidazole susceptibility, 2 linezolid-resistant isolates and 2 moxifloxacin-resistant isolates. We evaluated the efficacy of two cadazolid dosing regimens (250 versus 750 mg/L twice daily for 7 days) to treat simulated CDI. Microflora populations, C. difficile total viable counts and spores, cytotoxin titres, possible emergence of cadazolid, linezolid or quinolone resistance, and antimicrobial concentrations were monitored throughout., Results: Cadazolid was active against all (including linezolid- and moxifloxacin-resistant) C. difficile strains (MIC90 0.125, range 0.03-0.25 mg/L). The cadazolid geometric mean MIC was 152-fold, 16-fold, 9-fold and 7-fold lower than those of moxifloxacin, linezolid, metronidazole and vancomycin, respectively. Both cadazolid dosing regimens rapidly reduced C. difficile viable counts and cytotoxin with no evidence of recurrence. Cadazolid levels persisted at 50-100-fold supra-MIC for 14 days post-dosing. Cadazolid inhibition of enumerated gut microflora was limited, with the exception of bifidobacteria; Bacteroides fragilis group and Lactobacillus spp. counts were unaffected. There was no evidence for selection of strains resistant to cadazolid, quinolones or linezolid., Conclusions: Cadazolid activity was greater than other tested antimicrobials against 100 C. difficile strains. Cadazolid effectively treated simulated CDI in a gut model, with limited impact on the enumerated gut microflora and no signs of recurrence or emergence of resistance within the experimental timeframe.

Objectives: Fidaxomicin reduces the risk of recurrent Clostridium difficile infection (CDI) compared with vancomycin. We investigated fidaxomicin primary or secondary treatment efficacy using a gut model., Methods: Four triple-stage chemostat gut models were inoculated with faeces. After clindamycin induction of CDI, fidaxomicin (200 mg/L twice daily), vancomycin (125 mg/L four times daily) or metronidazole (9.3 mg/L three times daily) was administered for 7 days. Following failure/CDI recurrence, fidaxomicin (200 mg/L twice daily, 7 days) was instilled. C. difficile (CD) total viable counts (TVC), spore counts (SP), toxin titres (CYT), gut bacteria counts and antimicrobial concentrations were measured throughout., Results: Fidaxomicin instillation reduced CD TVC/SP and CYT below the limit of detection (LOD) after 2 and 4 days, respectively, with no CDI recurrence. Metronidazole instillation failed to decrease CD TVC or CYT. Vancomycin instillation reduced CD TVC and CYT to LOD by day 4, but SP persisted. Recurrence occurred 13 days after vancomycin instillation; subsequent fidaxomicin instillation reduced CD TVC/SP/CYT below the LOD from day 2. CD was isolated sporadically, with no evidence of spore recrudescence or toxin production. Fidaxomicin had a minimal effect on the microflora, except for bifidobacteria. Fidaxomicin was detected for at least 21 days post-instillation, whereas other antimicrobials were undetectable beyond ∼4 days., Conclusions: Fidaxomicin successfully treated simulated primary and recurrent CDI. Fidaxomicin was superior to metronidazole in reducing CD TVC and SP, and superior to vancomycin in reducing SP without recurrence of vegetative cell growth. Fidaxomicin, but not vancomycin or metronidazole, persisted in the gut model for >20 days after instillation.

No study to date has compared multilocus variable-number tandem-repeat analysis (MLVA) and whole-genome sequencing (WGS) in an investigation of the transmission of Clostridium difficile infection. Isolates from 61 adults with ongoing and/or recurrent C. difficile infections and 17 asymptomatic carriage episodes in children (201 samples), as well as from 61 suspected outbreaks affecting 2 to 41 patients in 31 hospitals in the United Kingdom (300 samples), underwent 7-locus MLVA and WGS in parallel. When the first and last samples from the same individual taken for a median (interquartile range [IQR]) of 63 days (43 to 105 days) apart were compared, the estimated rates of the evolution of single nucleotide variants (SNVs), summed tandem-repeat differences (STRDs), and locus variants (LVs) were 0.79 (95% confidence interval [CI], 0.00 to 1.75), 1.63 (95% CI, 0.00 to 3.59), and 1.21 (95% CI, 0.00 to 2.67)/called genome/year, respectively. Differences of >2 SNVs and >10 STRDs have been used to exclude direct case-to-case transmission. With the first serial sample per individual being used to assess discriminatory power, across all pairs of samples sharing a PCR ribotype, 192/283 (68%) differed by >10 STRDs and 217/283 (77%) by >2 SNVs. Among all pairs of cases from the same suspected outbreak, 1,190/1,488 (80%) pairs had concordant results using >2 SNVs and >10 STRDs to exclude transmission. For the discordant pairs, 229 (15%) had ≥2 SNVs but ≤10 STRDs, and 69 (5%) had ≤2 SNVs but ≥10 STRDs. Discordant pairs had higher numbers of LVs than concordant pairs, supporting the more diverse measure in each type of discordant pair. Conclusions on whether the potential outbreaks were confirmed were concordant in 58/61 (95%) investigations. Overall findings using MLVA and WGS were very similar despite the fact that they analyzed different parts of the bacterial genome. With improvements in WGS technology, it is likely that MLVA locus data will be available from WGS in the near future.

Background: The control of Clostridium difficile infection is a major international healthcare priority, hindered by a limited understanding of transmission epidemiology for these bacteria. However, transmission studies of bacterial pathogens are rapidly being transformed by the advent of next generation sequencing., Results: Here we sequence whole C. difficile genomes from 486 cases arising over four years in Oxfordshire. We show that we can estimate the times back to common ancestors of bacterial lineages with sufficient resolution to distinguish whether direct transmission is plausible or not. Time depths were inferred using a within-host evolutionary rate that we estimated at 1.4 mutations per genome per year based on serially isolated genomes. The subset of plausible transmissions was found to be highly associated with pairs of patients sharing time and space in hospital. Conversely, the large majority of pairs of genomes matched by conventional typing and isolated from patients within a month of each other were too distantly related to be direct transmissions., Conclusions: Our results confirm that nosocomial transmission between symptomatic C. difficile cases contributes far less to current rates of infection than has been widely assumed, which clarifies the importance of future research into other transmission routes, such as from asymptomatic carriers. With the costs of DNA sequencing rapidly falling and its use becoming more and more widespread, genomics will revolutionize our understanding of the transmission of bacterial pathogens.

With the frequency of cases of Clostridium difficile infection (CDI) increasing in many developed countries, accurate and reliable laboratory diagnosis of CDI is more important than ever. However, the diagnosis of CDI has been handicapped by the existence of two reference standards, one of which detects C. difficile toxin (cytotoxin assay) and the other only toxigenic strains (cytotoxigenic culture). Being relatively slow and laborious to perform, these reference methods were largely abandoned as routine diagnostic methods for toxin detection in favour of stand-alone rapid enzyme immunoassays (EIAs), which have suboptimal sensitivity and specificity. The management of CDI is undermined by high rates of both false-positive and false-negative test results. More recently developed nucleic acid amplification tests (NAATs) for toxin gene detection offer improved sensitivity over immunoassays, but fail to discriminate between CDI and asymptomatic colonization with C. difficile, and have clear drawbacks as stand-alone diagnostic tests. Two-step or three-step diagnostic algorithms have been proposed as a solution. In a large study of the effectiveness of currently available tests, a diagnostic algorithm was developed that combines available tests to more effectively distinguish patients with CDI from uninfected patients. This two-test protocol, which is now used in National Health Service laboratories in England, comprises an EIA for glutamate dehydrogenase detection or NAATs for toxin gene detection, followed by a relatively sensitive toxin EIA. This algorithm also identifies 'potential C. difficile excretors', individuals with diarrhoeal samples that contain C. difficile but without demonstrable toxin, who may be a source of transmission of C. difficile to susceptible patients., (© 2012 The Author Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.)

The morbidity and mortality associated with Clostridium difficile ribotype 078 were examined by comparison with other known outbreak strains. A healthcare interaction within eight weeks of a positive specimen significantly increased the likelihood of ribotype 078 compared with ribotype 027. Individuals with ribotype 078 also tended to come from community sources, have a hospital stay post specimen similar to ribotype 027 and a lower 30-day mortality, but these differences were not statistically significant. This study generates several hypotheses and a methodological platform to explore this unique profile., (Copyright © 2012 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Objectives: We previously demonstrated that 7 days of oritavancin instillation effectively treats Clostridium difficile infection (CDI) in a human gut model. Oritavancin may be more effective than vancomycin due to apparently increased activity against spores. We compared the efficacy of shortened dosing duration (4 days) of oritavancin and vancomycin for CDI treatment using the gut model., Methods: Clindamycin induced CDI in two triple-stage chemostat gut models primed with pooled human faeces and C. difficile ribotype 027 spores. Oritavancin (64 mg/L twice daily) or vancomycin (125 mg/L four times daily) was instilled for 4 days and the effects on C. difficile proliferation and toxin production, and gut microflora were determined., Results: Both oritavancin and vancomycin reduced toxin to undetectable levels. Recurrent C. difficile germination occurred 20 days after vancomycin instillation, with high-level toxin production. Oritavancin reduced C. difficile counts to around the detection limit for the remainder of the experiment, with spores undetectable from day 1 of instillation. Toxin production was reduced to below detectable levels, but was sporadically seen later, despite no evidence of germination. Both oritavancin and vancomycin instillation led to only modest effects on gut microflora., Conclusions: Shortened courses of oritavancin and vancomycin effectively treated CDI in a human gut model, but evidence of recurrence was observed following vancomycin instillation. Oritavancin exposure inhibited the recovery of C. difficile spores, as previously described. Shortened antibiotic exposure minimizes disruption to the gut microflora. These data indicate the possible value of a 4 day oritavancin dosing regimen for CDI treatment.

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.

PubMed, EMBASE and conference abstracts were reviewed systematically to determine the clinical and economic burden associated with Clostridium difficile infection (CDI) acquired and treated in European healthcare facilities. Inclusion criteria were: published in the English language between 2000 and 2010, and study population of at least 20 patients with documented CDI acquired/treated in European healthcare facilities. Data collection was completed by three unblinded reviewers using the Cochrane Handbook and PRISMA statement. The primary outcomes were mortality, recurrence, length of hospital stay (LOS) and cost related to CDI. In total, 1138 primary articles and conference abstracts were identified, and this was narrowed to 39 and 30 studies, respectively. Data were available from 14 countries, with 47% of studies from UK institutions. CDI mortality at 30 days ranged from 2% (France) to 42% (UK). Mortality rates more than doubled from 1999 to 2004, and continued to rise until 2007 when reductions were noted in the UK. Recurrent CDI varied from 1% (France) to 36% (Ireland); however, recurrence definitions varied between studies. Median LOS ranged from eight days (Belgium) to 27 days (UK). The incremental cost of CDI was £4577 in Ireland and £8843 in Germany, after standardization to 2010 prices. Country-specific estimates, weighted by sample size, ranged from 2.8% to 29.8% for 30-day mortality and from 16 to 37 days for LOS. CDI burden in Europe was most commonly described using 30-day mortality, recurrence, LOS and cost data. The continued spread of CDI and resultant healthcare burden underscores the need for judicious use of antibiotics., (Copyright © 2012 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

In the past decade, the incidence of Clostridium difficile infections (CDI) with a more severe course has increased in Europe and North America. Assays that are capable of rapidly diagnosing CDI are essential. Two real-time PCRs (LUMC and LvI) targeting C. difficile toxin genes (tcdB, and tcdA and tcdB, respectively) were compared with the BD GeneOhm PCR (targeting the tcdB gene), using cytotoxigenic culture as a gold standard. In addition, a real-time PCR targeting the tcdC frameshift mutation at position 117 (Δ117 PCR) was evaluated for detecting toxigenic C. difficile and the presence of PCR ribotype 027 in stool samples. In total, 526 diarrheal samples were prospectively collected and included in the study. Compared with those for cytotoxigenic culture, sensitivity, specificity, positive predicted value (PPV), and negative predicted value (NPV) were for PCR LUMC 96.0%, 88.0%, 66.0%, and 98.9%, for PCR LvI 100.0%, 89.4%, 69.7%, and 100.0%, for PCR Δ117 98.0%, 90.7%, 71.9%, and 99.5%, and for PCR BD GeneOhm 88.3%, 96.9%, 86.5%, and 97.4%. Compared to those with feces samples cultured positive for C. difficile type 027, the sensitivity, specificity, PPV, and NPV of the Δ117 PCR were 95.2%, 96.2%, 87.0%, and 98.7%. We conclude that all real-time PCRs can be applied as a first screening test in an algorithm for diagnosing CDI. However, the low PPVs hinder the use of the assays as stand-alone tests. Furthermore, the Δ117 PCR may provide valuable information for minimizing the spread of the epidemic C. difficile PCR ribotype 027.

Totals of 102 and 56 Clostridium difficile type 078 strains of human and porcine origins, respectively, from four European countries were investigated by an optimized multilocus variable-number tandem-repeat analysis (MLVA) and for tetracycline susceptibility. Eighty-five percent of all isolates were genetically related, irrespective of human or porcine origin. Human strains were significantly more resistant to tetracycline than porcine strains. All tetracycline-resistant strains contained the Tn916-like transposon harboring the tet(M) gene. We conclude that strains from human and porcine origins are genetically related, irrespective of the country of origin. This may reflect a lack of diversity and/or common source.

Clostridium difficile is an important healthcare-associated pathogen. Hypervirulent strains such as those belonging to ribotype 027 have been widely reported in recent years. A second strain associated with hypervirulence is ribotype 078 and the prevalence of Clostridium difficile infection (CDI) due to this ribotype appears to be increasing. This report describes an outbreak, in which 15cases of CDI due to ribotype 078 were detected in an Irish hospital and from a nursing home in the hospital's catchment area. C. difficile ribotype 078 accounted for 15% of total isolates submitted for ribotyping. The average age of patients with CDI due to ribotype 078 was 76 years. Forty-six percent of patients experienced recurrence of symptoms within eight weeks of diagnosis and CDI was felt to have directly contributed to five of the eight deaths. Use of enhanced DNA fingerprinting identified clusters within the 15 cases and suggested hitherto unrecognised links between some patients with CDI. Such approaches offer the promise to delineate common sources and transmission routes for C. difficile., (Copyright 2010 The Hospital Infection Society. Published by Elsevier Ltd. All rights reserved.)

The epidemiology of Clostridium difficile infection (CDI) has changed dramatically during this millennium. Infection rates have increased markedly in most countries with detailed surveillance data. There have been clear changes in the clinical presentation, response to treatment, and outcome of CDI. These changes have been driven to a major degree by the emergence and epidemic spread of a novel strain, known as PCR ribotype 027 (sometimes referred to as BI/NAP1/027). We review the evidence for the changing epidemiology, clinical virulence and outcome of treatment of CDI, and the similarities and differences between data from various countries and continents. Community-acquired CDI has also emerged, although the evidence for this as a distinct new entity is less clear. There are new data on the etiology of and potential risk factors for CDI; controversial issues include specific antimicrobial agents, gastric acid suppressants, potential animal and food sources of C. difficile, and the effect of the use of alcohol-based hand hygiene agents.

The aim of the present systematic review was to evaluate the available evidence on laboratory diagnosis of CDI and to formulate recommendations to optimize CDI testing. In comparison with cell culture cytotoxicity assay (CCA) and toxigenic culture (TC) of stools, we analyzed the test characteristics of 13 commercial available enzyme immunoasssays (EIA) detecting toxins A and/or B, 4 EIAs detecting Clostridium difficile glutamate dehydrogenase (GDH), and a real-time PCR for C. difficile toxin B gene. In comparison with CCA and TCA and assuming a prevalence of CDI of 5%, PPV and NPV varied between 0.28-0.77, 0.12-0.65 and 0.98-1.00, 0.97-1.00, respectively. Only if the tests were performed in a population with a CDI prevalence of 50 percent, would PPVs be acceptable (ranging from 0.71 to 1.00).To overcome the problem of a low PPV, we propose a two step approach, with a second test or a reference method in case of a positive first test. Further reducing the number of false negative results would require either retesting of all subjects with a negative first test, or re-testing all subjects with a negative second test, after an initially positive test. This approach resulted in non-significant improvements, and emphasizes the need for better diagnostic tests. Further studies to validate the applicability of two-step testing, including assessment of clinical features, are required.

To determine how best to decontaminate the hospital environment of Clostridium difficile, we carried out a cross-over study on two elderly medicine wards to determine whether cleaning with a hypochlorite disinfectant was better than using neutral detergent in reducing the incidence of C. difficile infection (CDI). We examined 1128 environmental samples in two years, 35% of which grew C. difficile. There was a significant decrease of CDI incidence on ward X, from 8.9 to 5.3 cases per 100 admissions (P<0.05) using hypochlorite, but there was no significant effect on ward Y. On ward X the incidence of CDI was significantly associated with the proportion of culture-positive environmental sites (P<0.05). On ward Y the only significant correlation between CDI and C. difficile culture-positive environmental sites was in patient side-rooms (r=0.41, P<0.05). The total daily defined doses of cefotaxime, cephradine and aminopenicillins were similar throughout the trial. These results provide some evidence that use of hypochlorite for environmental cleaning may significantly reduce incidence of CDI, but emphasize the potential for confounding factors.

We used three different DNA fingerprinting techniques and clindamycin susceptibility testing to confirm that Clostridium difficile PCR ribotype I isolates can be divided into two subclones. This observation may permit a better understanding of the epidemiology and pathogenicity of C. difficile infection.

Clostridium perfringens has been reported as the cause of up to 15% of cases of antibiotic-associated diarrhoea (AAD) and may be diagnosed by detection of enterotoxin (CPEnt) in faeces. The performance of a commercial ELISA method for CPEnt, with culture and PCR methods to confirm the presence of enterotoxigenic C. perfringens, was evaluated in 200 consecutive specimens from patients with clinical details suggestive of AAD: 8% of the specimens were positive for CPEnt, 16% were positive for C. difficile cytotoxin and 2% gave positive test results for both C. perfringens and C. difficile toxins. Culture and PCR results confirmed the majority of ELISA results, although 2 (12.5%) reactive specimens were only weakly positive. C. perfringens is a potentially important cause of infective AAD and can be detected with the C. perfringens enterotoxin ELISA kit, although weak positive results should be considered with caution.