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Two recently completed phase 3 trials (003 and 004) showed fidaxomicin to be noninferior to vancomycin for curing Clostridium difficile infection (CDI) and superior for reducing CDI recurrences. In both studies, adults with active CDI were randomized to receive blinded fidaxomicin 200 mg twice daily or vancomycin 125 mg 4 times a day for 10 days. Post hoc exploratory intent-to-treat (ITT) time-to-event analyses were undertaken on the combined study 003 and 004 data, using fixed-effects meta-analysis and Cox regression models. ITT analysis of the combined 003/004 data for 1164 patients showed that fidaxomicin reduced persistent diarrhea, recurrence, or death by 40% (95% confidence interval [CI], 26%—51%; P < .0001) compared with vancomycin through day 40. A 37% (95% CI, 2%—60%; P = .037) reduction in persistent diarrhea or death was evident through day 12 (heterogeneity P = .50 vs 13—40 days), driven by 7 (1.2%) fidaxomicin versus 17 (2.9%) vancomycin deaths at <12 days. Low albumin level, low eosinophil count, and CDI treatment preenrollment were risk factors for persistent diarrhea or death at 12 days, and CDI in the previous 3 months was a risk factor for recurrence (all P < .01). Fidaxomicin has the potential to substantially improve outcomes from CDI.

We determined the complete genome sequence of Clostridium difficile strain 630, a virulent and multidrug-resistant strain. Our analysis indicates that a large proportion (11%) of the genome consists of mobile genetic elements, mainly in the form of conjugative transposons. These mobile elements are putatively responsible for the acquisition by C. difficile of an extensive array of genes involved in antimicrobial resistance, virulence, host interaction and the production of surface structures. The metabolic capabilities encoded in the genome show multiple adaptations for survival and growth within the gut environment. The extreme genome variability was confirmed by whole-genome microarray analysis; it may reflect the organism's niche in the gut and should provide information on the evolution of virulence in this organism.

The strict anaerobe Clostridium difficile is the most common cause of nosocomial diarrhea, and the oxygen-resistant spores that it forms have a central role in the infectious cycle. The late stages of sporulation require the mother cell regulatory protein σK. In Bacillus subtilis, the onset of σK activity requires both excision of a prophage-like element (skinBs) inserted in the sigK gene and proteolytical removal of an inhibitory pro-sequence. Importantly, the rearrangement is restricted to the mother cell because the skinBs recombinase is produced specifically in this cell. In C. difficile, σK lacks a pro-sequence but a skinCd element is present. The product of the skinCd gene CD1231 shares similarity with large serine recombinases. We show that CD1231 is necessary for sporulation and skinCd excision. However, contrary to B. subtilis, expression of CD1231 is observed in vegetative cells and in both sporangial compartments. Nevertheless, we show that skinCd excision is under the control of mother cell regulatory proteins σE and SpoIIID. We then demonstrate that σE and SpoIIID control the expression of the skinCd gene CD1234, and that this gene is required for sporulation and skinCd excision. CD1231 and CD1234 appear to interact and both proteins are required for skinCd excision while only CD1231 is necessary for skinCd integration. Thus, CD1234 is a recombination directionality factor that delays and restricts skinCd excision to the terminal mother cell. Finally, while the skinCd element is not essential for sporulation, deletion of skinCd results in premature activity of σK and in spores with altered surface layers. Thus, skinCd excision is a key element controlling the onset of σK activity and the fidelity of spore development.

Key Points Disease that is associated with infection by Clostridium difficile represents an urgent public health threat. The severity of C. difficile infection is determined by strain virulence, interactions with intestinal commensal microbial communities, and the host immune response to damage of the intestinal epithelium that is induced by C. difficile . The ability to sporulate and germinate is essential to C. difficile virulence. Hundreds of genes that are involved in sporulation and germination have been identified as well as a bile acid receptor that induces germination. C. difficile secretes toxin proteins that are internalized by host cells through receptor-mediated endocytosis and cause disruption to cytoskeletal architecture, which leads to cell death. Toxin-mediated cell death results in the loss of intestinal barrier integrity and the translocation of bacteria into underlying tissues. The intestinal microbiota provides colonization resistance against C. difficile infection. Commensal bacteria that are capable of converting primary bile acids to secondary bile acids inhibit the growth of C. difficile by depriving C. difficile spores of an important germinant and by increasing the concentration of secondary bile acids in the intestinal lumen, which are toxic to the vegetative form of C. difficile . Toxin-mediated damage to the epithelium activates the host inflammatory immune response. The role of the immune system is to limit epithelial damage and the dissemination of intestinal bacteria into the circulation. However, an overly robust inflammatory response can be damaging to the host and contribute to disease pathology. Treating infection with Clostridium difficile and post-antibiotic disease can be difficult. In this Review, Abt, McKenney and Pamer show how insights into spore germination, virulence and interactions with the host and microbiota can help to combat this pathogen. Clostridium difficile is a major cause of intestinal infection and diarrhoea in individuals following antibiotic treatment. Recent studies have begun to elucidate the mechanisms that induce spore formation and germination and have determined the roles of C. difficile toxins in disease pathogenesis. Exciting progress has also been made in defining the role of the microbiome, specific commensal bacterial species and host immunity in defence against infection with C. difficile . This Review will summarize the recent discoveries and developments in our understanding of C. difficile infection and pathogenesis.

Recurrence of Clostridium difficile infection (CDI) occurs in approximately 25% of successfully treated patients. Two phase 3 randomized, double-blind trials were conducted at 154 sites in the United States, Canada, and Europe to compare fidaxomicin vs vancomycin in treating CDI. Patients with CDI received fidaxomicin 200 mg twice daily or vancomycin 125 mg 4 times daily for 10 days. The primary end point was clinical cure of CDI at end of treatment, and a secondary end point was recurrence during the 28 days following clinical cure. In all, 1164 subjects were enrolled, of which a subgroup of 128 in the per-protocol population had another recent episode of CDI prior to the CDI diagnosis at study enrollment. In the analysis of this subgroup, initial response to therapy was similar for both drugs (>90% cure). However, recurrence within 28 days occurred in 35.5% of patients treated with vancomycin and 19.7% of patients treated with fidaxomicin (-15.8% difference; 95% confidence interval, -30.4% to -0.3%; P = .045). Early recurrence (within 14 days) was reported in 27% of patients treated with vancomycin and 8% of patients treated with fidaxomicin (P = .003). In patients with a first recurrence of CDI, fidaxomicin was similar to vancomycin in achieving a clinical response at end of therapy but superior in preventing a second recurrence within 28 days. Clinical Trials Registration. NCT00314951 and NCT00468728.

The microflora-sparing properties of fidaxomicin were examined during the conduct of a randomized clinical trial comparing vancomycin 125 mg 4 times per day versus fidaxomicin 200 mg twice per day for 10 days as treatment of Clostridium difficile infection (CDI). Fecal samples were obtained from 89 patients (45 received fidaxomicin, and 44 received vancomycin) at study entry and on days 4, 10, 14, 21, 28, and 38 for quantitative cultures for C. difficile and cytotoxin B fecal filtrate concentrations. Additionally, samples from 10 patients, each receiving vancomycin or fidaxomicin, and 10 samples from healthy controls were analyzed by quantitative real-time polymerase chain reaction with multiple group-specific primers to evaluate the impact of antibiotic treatment on the microbiome. Compared with controls, patients with CDI at study entry had counts of major microbiome components that were 2—3-log 10 colony-forming units (CFU)/g lower. In patients with CDI, fidaxomicin allowed the major components to persist, whereas vancomycin was associated with a further 2—4-log 10 CFU reduction of Bacteroides/Prevotella group organisms, which persisted to day 28 of the study, and shorter term and temporary suppression of both Clostridium coccoides and Clostridium leptum group organisms. In the posttreatment period, C. difficile counts similarly persisted in both study populations, but reappearance of toxin in fecal filtrates was observed in 28% of vancomycin-treated patient samples (29 of 94), compared with 14% of fidaxomicin-treated patient samples (13 of 91; P = .03). Similarly, 23% of vancomycin-treated patients (10 of 44) and 11% of fidaxomicin-treated patients (5 of 44) had recurrence of CDI. Whereas vancomycin and fidaxomicin are equally effective in resolving CDI symptoms, preservation of the microflora by fidaxomicin is associated with a lower likelihood of CDI recurrence. Clinical Trials Registration. NTC00314951.

The dramatic changes in the epidemiology of Clostridium difficile infection (CDI) during recent years, with increases in incidence and severity of disease in several countries, have made CDI a global public health challenge. Increases in CDI incidence have been largely attributed to the emergence of a previously rare and more virulent strain, BI/NAP1/027. Increased toxin production and high-level resistance to fluoroquinolones have made this strain a very successful pathogen in healthcare settings. In addition, populations previously thought to be at low risk are now being identified as having severe CDI. Recent genetic analysis suggests that C. difficile has a highly fluid genome with multiple mechanisms to modify its content and functionality, which can make C. difficile adaptable to environmental changes and potentially lead to the emergence of more virulent strains. In the face of these changes in the epidemiology and microbiology of CDI, surveillance systems are necessary to monitor trends and inform public health actions.

Predicting dynamics of host-microbial ecosystems is crucial for the rational design of bacteriotherapies. We present MDSINE, a suite of algorithms for inferring dynamical systems models from microbiome time-series data and predicting temporal behaviors. Using simulated data, we demonstrate that MDSINE significantly outperforms the existing inference method. We then show MDSINE’s utility on two new gnotobiotic mice datasets, investigating infection with Clostridium difficile and an immune-modulatory probiotic. Using these datasets, we demonstrate new capabilities, including accurate forecasting of microbial dynamics, prediction of stable sub-communities that inhibit pathogen growth, and identification of bacteria most crucial to community integrity in response to perturbations.

Background. An epidemic strain of Clostridium difficile designated by restriction endonuclease analysis (REA) as group BI has caused multiple outbreaks of severe C. difficile infection (CDI). The treatment response of patients infected with this strain is uncertain. Methods. Clostridium difficile isolates were collected from 2 phase 3 clinical trials comparing fidaxomicin to vancomycin and typed using REA. Clinical cure and recurrence outcomes were analyzed by strain type of the infecting organism, BI and non-BI, using both univariate and multivariate analyses. Results. From 999 patients, 719 isolates were available for typing (356 fidaxomicin treated and 363 vancomycin treated). BI was the most common REA group (34% of isolates). Patients infected with BI had lower cure rates (86.6%; 214 of 247) than those infected with non-BI strains (94.3%; 445 of 472) (P < .001). The cure rate difference between the BI and non-BI patients was significant for both vancomycin (P = .02) and fidaxomicin (P = .007). BI patients had a recurrence rate of 27.4% (51 of 186), compared with a recurrence rate of 16.6% (66 of 397) in non-BI patients (P = .002). By multivariate analysis, BI infection was statistically significant as a risk factor for reduced cure (odds ratio [OR], 0.48; 95% confidence interval [CI], .27—.85; P = .030) and for increased recurrence (OR, 1.57; 95% CI, 1.01—2.45; P = .046). Conclusions. The clinical cure rate of patients infected with the epidemic BI C. difficile strain is lower than the cure rate of those infected with non-BI strains whether treated with fidaxomicin or vancomycin. Similarly, the CDI recurrence rate is increased in patients with the BI strain compared with patients with other C. difficile strains.

Clostridium difficile toxins revisited Clostridium difficile , the most common cause of infectious diarrhoea in hospitals in Europe and North America, produces two toxins. Their relative importance has been widely debated, and although animal studies had indicated that purified toxin A alone can induce most of the pathology observed in C. difficile infections, a recent Nature paper ( http://go.nature.com/oh6un5 ) suggested that the other toxin, toxin B, was the main cause of disease symptoms. Now a new study, involving C. difficile strains producing either toxin A or toxin B alone and — for the first time — a double-mutant strain producing neither, demonstrates that both toxins are important for disease, and need to be considered for diagnosis and treatment. Clostridium difficile , an important nosocomial pathogen, produces two toxins. Studies with purified toxins have indicated that only toxin A is important for pathogenesis, but recently it has been suggested that toxin B causes the majority of the disease symptoms in the context of a bacterial infection. These authors demonstrate that both toxins are important for disease and will need to be considered for diagnosis and treatment. Clostridium difficile infection is the leading cause of healthcare-associated diarrhoea in Europe and North America 1 , 2 . During infection, C. difficile produces two key virulence determinants, toxin A and toxin B. Experiments with purified toxins have indicated that toxin A alone is able to evoke the symptoms of C. difficile infection, but toxin B is unable to do so unless it is mixed with toxin A or there is prior damage to the gut mucosa 3 . However, a recent study indicated that toxin B is essential for C. difficile virulence and that a strain producing toxin A alone was avirulent 4 . This creates a paradox over the individual importance of toxin A and toxin B. Here we show that isogenic mutants of C. difficile producing either toxin A or toxin B alone can cause fulminant disease in the hamster model of infection. By using a gene knockout system 5 , 6 to inactivate the toxin genes permanently, we found that C. difficile producing either one or both toxins showed cytotoxic activity in vitro that translated directly into virulence in vivo . Furthermore, by constructing the first ever double-mutant strain of C. difficile, in which both toxin genes were inactivated, we were able to completely attenuate virulence. Our findings re-establish the importance of both toxin A and toxin B and highlight the need to continue to consider both toxins in the development of diagnostic tests and effective countermeasures against C. difficile .