Background: Carbapenemase-producing enterobacteriaceae (CPE) are a major threat for severely ill patients. However, only limited data on the epidemiology and on evidence-based infection prevention and control measures are available. The aim of this study was to investigate the epidemiology of patients with CPE, characterizing the CPE isolates by their resistance mechanisms and genetic similarity, to explore risk factors for their acquisition, and to evaluate the effectiveness of the current CPE infection control measures. Methods: A retrospective case-control study was performed using data from 2011 to 2016 in a 1800-bed academic hospital in Central Europe, where risk-based screening at patients´ admission is performed. Carbapenem resistance mechanisms of all carbapenem resistant enterobacteriaceae from patients admitted during this period were investigated. Clinical data of the CPE-positive patients were analysed and compared to a matched control group (case-control ratio of 1:3). We performed univariate and multivariate statistical analysis to identify risk factors for CPE acquisition. Results: Of 621,623 admitted patients in the study period, 75 patients with carriage of carbapenem resistant enterobacteriaceae were included (0.12/1000 admittances). Carbapenemase-encoding genes were detected in 77.3% (58/75) of patients with carbapenem-resistant enterobacteriaceae. The enzyme blaOXA-48 was found in 34.5% (20/58), blaKPC in 29.3% (17/58), blaNDM enzymes in 20.7% (12/58) and blaVIM in 8.6% (5/58) of the isolates. The overall mortality among CPE patients was 25.9% (15/58) and attributable mortality of CPE was 53.3% (8/15). Multivariate analysis revealed four risk factors to be independent predictors of CPE carriage: the length of hospital admission > 20 days (AOR: 4.9, 95% CI: 1.4–15.5; P < 0.001), hospital admission within the previous year (AOR: 22.3, 95% CI: 3.9–88.4; P < 0.001), exposure to a healthcare facility in a country with high or unknown carbapenem-resistant enterobacteriaceae prevalence 3 months before admission (AOR: 11.8, 95% CI: 2.2–63.2; P < 0.01) and the use of antibiotics longer than 10 days (AOR: 5.2, 95% CI: 1.4–35.9; P < 0.05). The current risk-based screening strategy at hospital admission could not identify 37 (63.8%) of the 58 CPE-positive patients. Epidemiological investigation and genotyping revealed that no outbreaks due to CPE occurred during this period. Conclusion: Overall, the CPE carriage rate in patients was very low, the attributable mortality, however, is alarming (53%). BlaOXA-48 and blaKPC were the main cause of carbapenem resistance in enterobacteriaceae. Although the strict application of standard infection control measures was effective for prevention of outbreaks in this setting, an enlarged risk based targeted screening strategy has to be implemented.

Staphylococcus lugdunensis is an emerging high-virulent pathogen causative of hospital-acquired infections. Biofilm formation is a complex pathogenic process that leads to well-established bacterial communities. There is a paucity of data on the composition of the biofilm matrix among S. lugdunensis strains. Here, twenty-two S. lugdunensis clinical isolates, mainly from orthopaedic infections but also from other clinical sources, were sub-grouped by ribotyping and dendrogram analysis. Biofilms were analysed by fluorimetric methods based on FITC-Wheat Germ Agglutinin, SYPRO Ruby and TOTO-1 dyes to detect exopolysaccharides, proteins and extracellular DNA (eDNA), respectively. Biofilm morphology was investigated under confocal laser scanning microscopy (CLSM). Isolates displayed intriguing diversities in biofilm mass and matrix composition. The content of exopolysaccharides was found to be to be strongly associated with the biofilm mass (R² = 0.882), while the content of proteins turned out to be weakly (R² = 0.465) and that of eDNA very weakly associated (R² = 0.202) to the biofilm mass.

As infection control faces new challenges from emerging, multidrug resistant strains of the yeast Candida auris, this study was conducted in order to examine the efficacy of hospital surface disinfectants and a new water disinfectant against C. auris biofilm forms. We tested four reference strains of C. auris (NCPF8971, NCPF8977, NCPF8984, DSM21092) and one C. albicans strain (ATCC10231) against disinfectants based on ethanol (ETH), quaternary ammonium (QAC), a combination of glutaraldehyde, quaternary ammonium and surfactant (ALD) and potassium peroxymonosulfate (PP) as well as 3.4% H₂O₂ and 4.25% H₂O₂ alone. In addition, a micellic-based formulation containing 17% v/v hydrogene peroxide disinfectant (mH₂O₂) was tested. The efficacy of the disinfectants was measured in a 96-well plate using tetrazolium salt reduction (XTT) and the log₁₀ reduction assay. ETH and QAC displayed in clinically recommended concentrations more than 5log₁₀ reduction and more than 80% XTT activity reduction for all of the Candida biofilms and planktonic cells. Only biofilms of C. auris NCPF8984 were additionally sensitive to all remaining disinfectants. All tested C. auris biofilms were sensitive to PP disinfectant and showed more than 5log₁₀ reduction. However, the XTT assay showed a reduction of less than 80% for the PP disinfectant, indicating the presence of active but non-culturable cells. The 25% mH₂O₂ (corresponding to 4.25% H₂O₂) killed Candida biofilms after 1 minute.

Background Prosthetic implant infections caused by Staphylococcus aureus and epidermidis are major challenges for early diagnosis and treatment owing to biofilm formation on the implant surface. Extracellular DNA (eDNA) is actively excreted from bacterial cells in biofilms, contributing to biofilm stability, and may offer promise in the detection or treatment of such infections. Questions/purposes (1) Does DNA structure change during biofilm formation? (2) Are there time-dependent differences in eDNA production during biofilm formation? (3) Is there differential eDNA production between clinical and control Staphylococcal isolates? (4) Is eDNA production correlated to biofilm thickness? Methods We investigated eDNA presence during biofilm formation in 60 clinical and 30 control isolates of S aureus and S epidermidis. The clinical isolates were isolated from patients with infections of orthopaedic prostheses and implants: 30 from infected hip prostheses and 30 from infected knee prostheses. The control isolates were taken from healthy volunteers who had not been exposed to antibiotics and a hospital environment during the previous 3 and 12 months, respectively. Control S epidermidis was isolated from the skin of the antecubital fossa, and control S aureus was isolated from the nares. For the biofilm experiments the following methods were used to detect eDNA: (1) fluorescent staining with 4′,6-diamidino-2-phenylindole (DAPI), (2) eDNA extraction using a commercial kit, and (3) confocal laser scanning microscopy for 24-hour biofilm observation using propidium iodide and concanavalin-A staining; TOTO^®-1 and SYTO^® 60 staining were used for observation and quantification of eDNA after 6 and 24 hours of biofilm formation. Additionally antibiotic resistance was described. Results eDNA production as observed by confocal laser scanning microscopy was greater in clinical isolates than controls (clinical isolates mean ± SD: 1.84% ± 1.31%; control mean ± SD: 1.17% ± 1.37%; p < 0.005) after 6 hours of biofilm formation. After 24 hours, the amount of eDNA was greater in biofilms of S epidermidis than in biofilms of S aureus (S aureus mean ± SD: 1.35% ± 2.0%; S epidermidis mean ± SD: 6.42% ± 10.6%; p < 0.05). Clinical isolates of S aureus and S epidermidis produced more eDNA than control isolates at 6 hours of biofilm formation. The extraction method also showed that clinical isolates produced substantially greater amounts of eDNA than controls. Conclusions S aureus and S epidermidis exhibit a differential production of DNA with time. Clinical isolates associated with implant infections produce greater amounts of eDNA than controls. Future research might focus on the diagnostic value of eDNA as a surrogate laboratory marker for biofilm formation in implant infections. Clinical relevance eDNA should be considered as a potential future diagnostic tool or even a possible target to modify biofilms for successful treatment of biofilm-associated infections.

Aims. Microbial colonization of silicone voice prostheses by bacteria andCandidaspecies limits the device lifetime of modern voice prostheses in laryngectomized patients. Thus, research focuses on biofilm inhibitive properties of novel materials, coatings, and surface enhancements. Goal of thisin vitrostudy was the evaluation of seven commonly used growth media to simulate growth of mixed oropharyngeal species as mesoscale biofilms on prosthetic silicone for future research purposes.Methods and Results. Yeast Peptone Dextrose medium (YPD), Yeast Nitrogen Base medium (YNB), M199 medium, Spider medium, RPMI 1640 medium, Tryptic Soy Broth (TSB), and Fetal Bovine Serum (FBS) were used to culture combined mixedCandidastrains and mixed bacterial-fungal compositions on silicone over the period of 22 days. The biofilm surface spread and the microscopic growth showed variations fromin vivobiofilms depending on the microbial composition and growth medium.Conclusion. YPD and FBS prove to support continuousin vitrogrowth of mixed bacterial-fungal oropharyngeal biofilms deposits over weeks as needed for longtermin vitrotesting with oropharyngeal biofilm compositions.Significance and Impact of Study. The study provides data on culture conditions for mixed multispecies biofilm compositions that can be used for future prosthesis designs.

In a retrospective study the association of the production of extracellular DNA (eDNA) in biofilms of clinical staphylococcal isolates from 60 patients with prosthetic joint infection (PJI) and the clinical outcome were investigated. Data from a previous study on eDNA production determined in 24-hour biofilms of staphylococcal isolates (Staphylococcus aureusn=30,Staphylococcus epidermidisn=30) was correlated with the patients’ clinical outcome after 3 and 12 months. Statistical analysis was performed using either the Spearman’s rank correlations test or the t-test. eDNA production ofS. epidermidisin 24-hour biofilms correlated with the patients’ outcome ‘not cured‘ after 12 months. ForS. aureusno such correlation was detected. Thus, eDNA may be a virulence factor ofS. epidermidis.Quantification of eDNA production as a surrogate marker for biofilm formation might be a potential predictive marker for the management of PJI.