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Genomic analysis of a diverse collection of Clostridioides difficile ribotype 078 isolates from Ireland and 9 countries in Europe provided evidence for complex regional and international patterns of dissemination that are not restricted to humans. These isolates are associated with C. difficile colonization and clinical illness in humans and pigs. [ABSTRACT FROM AUTHOR] more...

ABSTRACT Insights into the interaction between phages and their bacterial hosts are crucial for the development of phage therapy. However, only one study has investigated global gene expression of Clostridioides (formerly Clostridium) difficile carrying prophage, and transcriptional reprogramming during lytic infection has not been studied. Here, we presented the isolation, propagation, and characterization of a newly discovered 35,109-bp phage, JD032, and investigated the global transcriptomes of both JD032 and C. difficile ribotype 078 (RT078) strain TW11 during JD032 infection. Transcriptome sequencing (RNA-seq) revealed the progressive replacement of bacterial host mRNA with phage transcripts. The expressed genes of JD032 were clustered into early, middle, and late temporal categories that were functionally similar. Specifically, a gene (JD032_orf016) involved in the lysis-lysogeny decision was identified as an early expression gene. Only 17.7% (668/3,781) of the host genes were differentially expressed, and more genes were downregulated than upregulated. The expression of genes involved in host macromolecular synthesis (DNA/RNA/proteins) was altered by JD032 at the level of transcription. In particular, the expression of the ropA operon was downregulated. Most noteworthy is that the gene expression of some antiphage systems, including CRISPR-Cas, restriction-modification, and toxin-antitoxin systems, was suppressed by JD032 during infection. In addition, bacterial sporulation, adhesion, and virulence factor genes were significantly downregulated. This study provides the first description of the interaction between anaerobic spore-forming bacteria and phages during lytic infection and highlights new aspects of C. difficile phage-host interactions. IMPORTANCE C. difficile is one of the most clinically significant intestinal pathogens. Although phages have been shown to effectively control C. difficile infection, the host responses to phage predation have not been fully studied. In this study, we reported the isolation and characterization of a new phage, JD032, and analyzed the global transcriptomic changes in the hypervirulent RT078 C. difficile strain, TW11, during phage JD032 infection. We found that bacterial host mRNA was progressively replaced with phage transcripts, three temporal categories of JD032 gene expression, the extensive interplay between phage-bacterium, antiphage-like responses of the host and phage evasion, and decreased expression of sporulation- and virulence-related genes of the host after phage infection. These findings confirmed the complexity of interactions between C. difficile and phages and suggest that phages undergoing a lytic cycle may also cause different phenotypes in hosts, similar to prophages, which may inspire phage therapy for the control of C. difficile. more...

• Almost 20% of 93 bat guano samples contain Clostridioides difficile (C. difficile). • C. difficile ribotypes 078, 056 and a new ribotype (designated CDB3; A + B+CDT-) were identified. • Bats do not seem to be a significant source for C. difficile dissemination in the environment. • Dissemination of known and new C. difficile ribotypes from bats is still possible. Bats are associated with the emergence of several mammalian diseases. Their sessional migration, and tendency to form large colonies in close proximity to human habitats enables effective intra- and inter-species transmission of pathogens. Clostridioides difficile is an important enteric pathogen in humans and animals; however, the source of its dissemination in the population is unknown. The purpose of this study was to determine the prevalence of C. difficile in bats, and to characterize C. difficile isolates. Feces (n = 93) was sampled from bats during their migration across Europe. Eighteen samples (19.4%) were positive for C. difficile ; ribotypes 078, 056, and a new ribotype CDB3 were identified. Clostridioides difficile ribotypes 078 and 056 are associated with human and animal diseases. The C. difficile prevalence and ribotypes in this study do not necessarily identify bats as a significant source, but more likely as an indicator of C. difficile perpetuation in the environment. [ABSTRACT FROM AUTHOR] more...

Clostridioides (C.) difficile are found in cows, pigs and poultry suggesting that this pathogen is present and more importantly animals could act as a reservoir, via food or environment, of human C. difficile infection. Molecular methods together with phenotypical characterisation bring integrated and important tools to describe diversity and nature of bacteria including C. difficile. Moreover, similar or identical C. difficile types are found in different farm animals. This study aimed to phenotypically characterise C. difficile isolates belonging to ribotype 078 and to identify differences such as growth and toxicity between porcine and bovine isolates. C. difficile isolates were assessed for the growth behaviour (turbidimetry), metabolic potential (Biolog AN) and toxin production (ELISA method) in vitro. The concentration of released either toxin A (TcdA) or toxin B (TcdB) varied greatly between the isolates tested; however, it did not differ between the porcine and bovine ribotypes. Also, the TcdA/TcdB ratio of the isolates did not show a difference either. The most common metabolised substrates were pyruvic acid followed by α-ketobutyric acid. The results show that both porcine and bovine C. difficile RT 078 share similar phenotypical characteristics including growth and production of toxins. The findings may help understand the virulence of C. difficile RT 078 in porcine and bovine species. [ABSTRACT FROM AUTHOR] more...

Abstract Antimicrobial susceptibility to 6 antimicrobial agents, PCR-ribotyping and molecular genetics of fluoroquinolone resistance was assessed in 70 toxigenic clinical isolates of C. difficile recovered from patients attended in a hospital in southern Spain with suspected Clostridium difficile infection. Moxifloxacin was the least active drug, mainly driven by the aminoacid substitution Thr82Ile in GyrA, while PCR-ribotype 078 was the most prevalent lineage identified and grouped several of the fluoroquinolone resistant isolates. Highlights • Antimicrobial susceptibility and molecular typing of C. difficile are performed. • Vancomycin, linezolid and tigecycline were the most active agents. • Ribotype 078 was the most prevalent (20%), followed by ribotype 014 (17%). • A high prevalence of fluorquinolone-resistant ribotype 078 is observed. • Thr82 to Ile in GyrA was found in the majority of the resistant isolates. [ABSTRACT FROM AUTHOR] more...

Clostridium difficile can cause neonatal enteritis, but has been isolated from both diseased and healthy pigs. C. difficile shedding by pigs is a potential source of zoonotic transmission to humans. The objectives of this study were to characterize the prevalence and genotype of C. difficile with respect to age and health status in the pig industry for the first time in Korea. Fecal samples of 910 pigs were analyzed for the presence of C. difficile . In total, 176 (19.3%) C. difficile strains were isolated. The prevalence was significantly higher ( p  < 0.01) in diarrheic samples (30.4%) than in non-diarrheic samples (13.3%). The prevalence was highest in diarrheic suckling piglets (53.6%), followed by diarrheic sows (40.0%) and non-diarrheic suckling piglets (34.0%). The toxin genes tcdA, tcdB, cdtA, and cdtB were all detected in 153 isolates (A + B + CDT + , 86.9%). Furthermore, the majority of toxigenic strains (86.5%) were identified as PCR ribotype 078, which is an important pathogen in both pigs and humans. These results indicated that C. difficile is a leading causative pathogen of neonatal diarrhea in piglets in Korea. [ABSTRACT FROM AUTHOR] more...

Pigs, particularly piglets, have been identified as reservoir hosts of Clostridium difficile. To examine the survival ability of this pathogen in pig feces-based manure compost, C. difficile spores, which were prepared to contain as few vegetative cells as possible, were artificially inoculated into pig feces and incubated at different temperatures. While C. difficile survived in the feces incubated at temperatures below 37 °C for over 30 days, cell numbers gradually decreased at thermophilic temperatures (over 55 °C; p < 0.05). Next, to clarify the prevalence of C. difficile in field manure compost, we isolated and characterized C. difficile from the final products of manure compost products of 14 pig farms. A total of 11 C. difficile strains were isolated from 5 of 14 (36% positive rate) samples tested. Of these 11 strains, 82% were toxigenic, with ribotype 078 being the most prevalent. Thus, the application of composted manure to land therefore poses a possible risk of C. difficile transfer to the food chain. [ABSTRACT FROM AUTHOR] more...

Clostridium difficile is a human and animal pathogen. Recently, the incidence of community-acquired C. difficile infection has increased, and many studies have indicated that C. difficile might be food-borne. The correlation between C. difficile infection in humans and in animals has been a topic of debate. The objective of this study was to determine the genetic relatedness of C. difficile from human and pigs in Taiwan. We investigated the molecular epidemiology of C. difficile in healthy humans and pigs from 2011 to 2015. The isolation rate of C. difficile from pigs in 13 commercial farms was 49% (100/204), and a high proportion of hypervirulent ( C. difficile carrying tcdA , tcdB , and cdtA/B genes and a 39-bp deletion in the tcdC gene) ribotype 078 lineage isolates (90%, 90/100; including 078, 126, 127, and 066-like isolates) were identified. In addition, the C. difficile ribotype 127 isolates from pigs typically exhibited moxifloxacin resistance (37/43; 86%). In healthy humans, the isolation rate was 4.3% (3/69), and all healthy human isolates were non-toxigenic. In particular, we compared the porcine isolates with two patient strains (ribotype 127) obtained from two hospitals in central Taiwan. The multilocus variable number tandem repeat analysis revealed a high genetic relatedness between ribotype 127 from patients and pigs. This study indicated that isolates of the ribotype 078 lineage, and especially ribotype 127, were widely distributed in pig farms and showed a high frequency of moxifloxacin resistance. The closely related ribotype 127 from patients and pigs may have had a common origin or low diversity. In conclusion, C. difficile ribotype 127 is a noteworthy pathogen in pigs and poses a potential public health threat. [ABSTRACT FROM AUTHOR] more...

Background: Research has indicated that the environment may play an important role in the transmission of meticillin-resistant Staphylococcus aureus (MRSA) and Clostridium difficile in healthcare facilities. Despite the significance of this finding, few data exist from longitudinal studies investigating MRSA and C. difficile contamination, concurrently, in both patient rooms and the general ward environment. The objectives of this study were to determine the prevalence of MRSA and C. difficile contamination in patient rooms and the ward environment and identify risk factors associated with a surface being contaminated with these pathogens. Methods: Environmental surfaces in patient rooms and the general environment in the medical and surgical wards of a community hospital were sampled six times over a 15 week period. Sterile electrostatic cloths were used for sampling and information pertaining to the surface sampled was recorded. MRSA isolates and C. difficile specimens were obtained from hospitalized patients. Enrichment culture was performed and spa typing or ribotyping was conducted for MRSA or C. difficile, respectively. Exact logistic regression models were constructed to examine risk factors associated with MRSA and C. difficile contamination. Results: Sixteen (41%) patient rooms had ⩾ 1 surfaces contaminated with MRSA and/or C. difficile. For 218 surfaces investigated, 3.2% and 6.4% were contaminated with MRSA or C. difficile, respectively. Regression models indicated that surfaces in rooms exposed to a C. difficile patient had significantly increased odds of being contaminated with C. difficile, compared to surfaces in unexposed patient rooms. Additionally, compared to plastic surfaces, cork surfaces had significantly increased odds of being contaminated with C. difficile. For 236 samples collected from the ward environment, MRSA and C. difficile were recovered from 2.5% and 5.9% of samples, respectively. Overall, the majority of MRSA and C. difficile strains were molecularly identified as spa type 2/t002 (84.6%, n = 11) and ribotype 078 (50%, n = 14), respectively. Conclusions: In patient rooms and the ward environment, specific materials and locations were identified as being contaminated with MRSA or C. difficile. These sites should be cleaned and disinfected with increased vigilance to help limit the transmission and dissemination of MRSA and C. difficile within the hospital. [ABSTRACT FROM AUTHOR] more...

Summary: 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 &y& Elsevier] more...

Abstract: Concern has been raised about the potential for Clostridium difficile to be a bovine and foodborne pathogen, yet limited study has been performed in cattle, and none in veal calves. This study evaluated the epidemiology and microbiology of C. difficile on one veal farm. Rectal swabs were obtained from calves within 48h of arrival and at one, 17 and 21 weeks later. Selective culture for C. difficile was performed. Isolates were characterized by PCR ribotyping and PCR for tcdA, tcdB and cdtA. Tetracycline resistance and resistance genes were investigated. Multivariable logistic regression models were constructed to determine the relationship between shedding of the bacterium and specific ribotypes and the independent variables: time of sampling and area of housing. Calves were twice more likely to test positive 1 week after arrival (51%) when compared to initial results (32%). Shedding at 17 and 21 weeks was significantly lower (2% at both samplings). Ribotype 078 was the most common. Twelve different ribotypes were present initially with only three ribotypes found subsequently. Seventy-six percent (40/53) of isolates initially recovered were tetracycline resistant compared to 93% (81/87) from 2nd sampling. Tetracycline resistance genes were detected in 24% (13/53) of isolates during 1st and in 55% (50/91) during 2nd sampling. The high prevalence of pathogenic C. difficile in veal calves could be of zoonotic concern. The low prevalence before slaughter may be of importance for the evaluation of foodborne risks. Oxytetracycline administration to calves may have an impact on prevalence of C. difficile colonization. [Copyright &y& Elsevier] more...

Abstract: Clostridium difficile is the primary known cause of antibiotic-associated diarrhea. Diarrheal disease in food animals due to C. difficile infection has been well documented. Recently, reports of C. difficile infections in patients with no known risk factors for disease have raised concern of community acquisition through food animals and food. In this study, multi-locus variable number tandem repeat analysis (MLVA) was performed on a collection of 97 C. difficile isolates of human, animal and food origin belonging to either the North American pulsed-field type (NAP) 1 or NAP7/NAP8. MLVA discriminated between NAP1 and NAP7/NAP8 populations. Three clusters of food, food animal and human NAP1 isolates were highly related by MLVA. These data suggest the possibility of either laboratory contamination or widespread distribution of clonal C. difficile populations. Community-associated NAP1 isolates were unrelated to NAP1 food and food animal isolates. Two MLVA loci were absent and 1 was invariant in all NAP7/NAP8 isolates. Therefore, MLVA discrimination was not sufficient to make assessments regarding the genetic associations among food, food animal and human isolates belonging to the NAP7/NAP8 pulsovar. Rigorous epidemiologic and laboratory investigations that employ highly discriminatory genotyping methods are necessary to compare C. difficile isolates from food and food animals to those from humans. [Copyright &y& Elsevier] more...

Summary: 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 &y& Elsevier] more...