Skin dysbiosis and loss of microbiome site specificity in critically ill patients.

An increasing amount of evidence has linked critical illness with dysbiotic microbiome signatures in different body sites. The disturbance of the indigenous microbiota structures has been further associated with disease severity and outcome and has been suggested to pose an additional risk for complications in intensive care units (ICUs), including hospital-acquired infections. A better understanding of the microbial dysbiosis in critical illness might thus help to develop strategies for the prevention of such complications. While most of the studies addressing microbiome changes in ICU patients have focused on the gut, the lung, or the oral cavity, little is known about the microbial communities on the skin of ICU patients. Since the skin is the outermost organ and the first immune barrier against pathogens, its microbiome might play an important role in the risk management for critically ill patients. This observational study characterizes the skin microbiome in ICU patients covering five different body sites at the time of admission. Our results show a profound dysbiosis on the skin of critically ill patients, which is characterized by a loss of site specificity and an overrepresentation of gut bacteria on all skin sites when compared to a healthy group. This study opens a new avenue for further investigations on the effect of skin dysbiosis in the ICU setting and points out the need of strategies for the management of dysbiosis in critically ill patients.IMPORTANCEUnbalanced gut microbiota in critically ill patients has been associated with poor outcome and complications during the intensive care unit (ICU) stay. Whether the disturbance of the microbial communities in these patients is extensive for other body sites, such as the skin, is largely unknown. The skin not only is the largest organ of the body but also serves as the first immune barrier against potential pathogens. This study characterized the skin microbiota on five different body sites in ICU patients at the time of admission. The observed disturbance of the bacterial communities might help to develop new strategies in the risk management of critically ill patients.

Staphylococcus aureus induces tolerance in human monocytes accompanied with expression changes of cell surface markers.

Exposure of human monocytes to lipopolysaccharide (LPS) or other pathogen-associated molecular pattern (PAMPs) induces a temporary insensitivity to subsequent LPS challenges, a cellular state called endotoxin tolerance (ET), associated with the pathogenesis of sepsis. In this study, we aimed to characterize the cellular state of human monocytes from healthy donors stimulated with Staphylococcus aureus in comparison to TLR2-specific ligands. We analyzed S. aureus induced gene expression changes after 2 and 24 hours by amplicon sequencing (RNA-AmpliSeq) and compared the pro-inflammatory response after 2 hours with the response in re-stimulation experiments. In parallel, glycoprotein expression changes in human monocytes after 24 hours of S. aureus stimulation were analyzed by proteomics and compared to stimulation experiments with TLR2 ligands Malp-2 and Pam3Cys and TLR4 ligand LPS. Finally, we analyzed peripheral blood monocytes of patients with S. aureus bloodstream infection for their ex vivo inflammatory responses towards S. aureus stimulation and their glycoprotein expression profiles. Our results demonstrate that monocytes from healthy donors stimulated with S. aureus and TLR ligands of Gram-positive bacteria entered the tolerant cell state after activation similar to LPS treatment. In particular reduced gene expression of pro-inflammatory cytokines (TNF, IL1ß) and chemokines (CCL20, CCL3, CCL4, CXCL2, CXCL3 and CXCL8) could be demonstrated. Glycoprotein expression changes in monocytes tolerized by the different TLR agonists were highly similar while S. aureus-stimulated monocytes shared some of the PAMP-induced changes but also exhibited a distinct expression profile. 11 glycoproteins (CD44, CD274, DSC2, ICAM1, LAMP3, LILRB1, PTGS2, SLC1A3, CR1, FGL2, and HP) were similarly up- or downregulated in all four comparisons in the tolerant cell state. Monocytes from patients with S. aureus bacteremia revealed preserved pro-inflammatory responsiveness to S. aureus stimulation ex vivo, expressed increased CD44 mRNA but no other glycoprotein of the tolerance signature was differentially expressed.

Comparative analysis of surface sanitization protocols on the bacterial community structures in the hospital environment.

OBJECTIVES: In hospital hygiene, it remains unclear to what extent surface contamination might represent a potential reservoir for nosocomial pathogens. This study investigates the effects of different sanitization strategies on the microbial structures and the ecological balance of the environmental microbiome in the clinical setting. METHODS: Three cleaning regimes (disinfectants, detergents, and probiotics) were applied subsequently in nine independent patient rooms at a neurological ward (Charité, Berlin). Weekly sampling procedures included three different environmental sites: floor, door handle, and sink. Characterization of the environmental microbiota and detection of antibiotic resistance genes (ARGs) were performed by 16S rRNA sequencing and multiplex Taq-Man qPCR assays, respectively. RESULTS: Our results showed a displacement of the intrinsic environmental microbiota after probiotic sanitization, which reached statistical significance in the sink samples (median 16S-rRNA copies = 138.3; IQR: 24.38-379.5) when compared to traditional disinfection measures (median 16S rRNA copies = 1343; IQR: 330.9-9479; p < 0.05). This effect was concomitant with a significant increase in the alpha-diversity metrics in both the floor (p < 0.001) and the sink samples (p < 0.01) during the probiotic strategy. We did not observe a sanitization-dependent change in relative pathogen abundance at any tested site, but there was a significant reduction in the total ARG counts in the sink samples during probiotic cleaning (mean ARGs/sample: 0.095 ± 0.067) when compared to the disinfection strategy (mean ARGs/sample: 0.386 ± 0.116; p < 0.01). DISCUSSION: The data presented in this study suggest that probiotic sanitization is an interesting strategy in hospital hygiene management to be further analyzed and validated in randomized clinical studies.

A simple sequence-based filtering method for the removal of contaminants in low-biomass 16S rRNA amplicon sequencing approaches.

Controlling for contaminant sequences in microbiome experiments involving low-biomass samples is a highly challenging task which still lacks of standardized protocols. Here we propose a simple sequence-based filtering method for 16S rRNA gene microbial profiling approaches, and validate its efficiency using mock community dilution series and environmental samples collected in a clinical setting.