A point-prevalence study on community and inpatient Clostridioides difficile infections (CDI): results from Combatting Bacterial Resistance in Europe CDI (COMBACTE-CDI), July to November 2018.

BackgroundThere is a paucity of data on community-based Clostridioides difficile infection (CDI) and how these compare with inpatient CDI.AimTo compare data on the populations with CDI in hospitals vs the community across 12 European countries.MethodsFor this point-prevalence study (July-November 2018), testing sites sent residual diagnostic material on sampling days to a coordinating laboratory for CDI testing and PCR ribotyping (n = 3,163). Information on whether CDI testing was requested at the original site was used to identify undiagnosed CDI. We used medical records to identify differences between healthcare settings in patient demographics and risk factors for detection of C. difficile with or without free toxin.ResultsThe CDI positivity rate was 4.4% (country range: 0-16.2) in hospital samples, and 1.3% (country range: 0-2.2%) in community samples. The highest prevalence of toxinotype IIIb (027, 181 and 176) was seen in eastern European countries (56%; 43/77), the region with the lowest testing rate (58%; 164/281). Different predisposing risk factors were observed (use of broad-spectrum penicillins in the community (OR: 8.09 (1.9-35.6), p = 0.01); fluoroquinolones/cephalosporins in hospitals (OR: 2.2 (1.2-4.3), p = 0.01; OR: 2.0 (1.1-3.7), p = 0.02)). Half of community CDI cases were undetected because of absence of clinical suspicion, accounting for three times more undiagnosed adults in the community compared with hospitals (ca 111,000 vs 37,000 cases/year in Europe).ConclusionThese findings support recommendations for improving diagnosis in patients presenting with diarrhoea in the community, to guide good practice to limit the spread of CDI.

Hemin availability induces coordinated DNA methylation and gene expression changes in Porphyromonas gingivalis.

Periodontal disease is a chronic inflammatory disease in which the oral pathogen Porphyromonas gingivalis plays an important role. Porphyromonas gingivalis expresses virulence determinants in response to higher hemin concentrations, but the underlying regulatory processes remain unclear. Bacterial DNA methylation has the potential to fulfil this mechanistic role. We characterized the methylome of P. gingivalis, and compared its variation to transcriptome changes in response to hemin availability. Porphyromonas gingivalis W50 was grown in chemostat continuous culture with excess or limited hemin, prior to whole-methylome and transcriptome profiling using Nanopore and Illumina RNA-Seq. DNA methylation was quantified for Dam/Dcm motifs and all-context N6-methyladenine (6mA) and 5-methylcytosine (5mC). Of all 1,992 genes analyzed, 161 and 268 were respectively over- and under-expressed with excess hemin. Notably, we detected differential DNA methylation signatures for the Dam "GATC" motif and both all-context 6mA and 5mC in response to hemin availability. Joint analyses identified a subset of coordinated changes in gene expression, 6mA, and 5mC methylation that target genes involved in lactate utilization and ABC transporters. The results identify altered methylation and expression responses to hemin availability in P. gingivalis, with insights into mechanisms regulating its virulence in periodontal disease. IMPORTANCE DNA methylation has important roles in bacteria, including in the regulation of transcription. Porphyromonas gingivalis, an oral pathogen in periodontitis, exhibits well-established gene expression changes in response to hemin availability. However, the regulatory processes underlying these effects remain unknown. We profiled the novel P. gingivalis epigenome, and assessed epigenetic and transcriptome variation under limited and excess hemin conditions. As expected, multiple gene expression changes were detected in response to limited and excess hemin that reflect health and disease, respectively. Notably, we also detected differential DNA methylation signatures for the Dam "GATC" motif and both all-context 6mA and 5mC in response to hemin. Joint analyses identified coordinated changes in gene expression, 6mA, and 5mC methylation that target genes involved in lactate utilization and ABC transporters. The results identify novel regulatory processes underlying the mechanism of hemin regulated gene expression in P. gingivalis, with phenotypic impacts on its virulence in periodontal disease.

Immunomodulatory streptococci that inhibit CXCL8 secretion and NFκB activation are common members of the oral microbiota.

Introduction. Oral tissues are generally homeostatic despite exposure to many potential inflammatory agents including the resident microbiota. This requires the balancing of inflammation by regulatory mechanisms and/or anti-inflammatory commensal bacteria. Thus, the levels of anti-inflammatory commensal bacteria in resident populations may be critical in maintaining this homeostatic balance.Hypothesis/Gap Statement. The incidence of immunosuppressive streptococci in the oral cavity is not well established. Determining the proportion of these organisms and the mechanisms involved may help to understand host-microbe homeostasis and inform development of probiotics or prebiotics in the maintenance of oral health.Aim. To determine the incidence and potential modes of action of immunosuppressive capacity in resident oral streptococci.Methodology. Supragingival plaque was collected from five healthy participants and supragingival and subgingival plaque from five with gingivitis. Twenty streptococci from each sample were co-cultured with epithelial cells±flagellin or LL-37. CXCL8 secretion was detected by ELISA, induction of cytotoxicity in human epithelial cells by lactate dehydrogenase release and NFκB-activation using a reporter cell line. Bacterial identification was achieved through partial 16S rRNA gene sequencing and next-generation sequencing.Results. CXCL8 secretion was inhibited by 94/300 isolates. Immunosuppressive isolates were detected in supragingival plaque from healthy (4/5) and gingivitis (4/5) samples, and in 2/5 subgingival (gingivitis) plaque samples. Most were Streptococcus mitis/oralis. Seventeen representative immunosuppressive isolates all inhibited NFκB activation. The immunosuppressive mechanism was strain specific, often mediated by ultra-violet light-labile factors, whilst bacterial viability was essential in certain species.Conclusion. Many streptococci isolated from plaque suppressed epithelial cell CXCL8 secretion, via inhibition of NFκB. This phenomenon may play an important role in oral host-microbe homeostasis.