The choice of effective biocides used for routine hospital practice should consider the role of disinfectants in the maintenance and development of local resistome and how they might affect antibiotic resistance gene transfer within the hospital microbial population. Currently, there is little understanding of how different biocides contribute to eDNA release that may contribute to gene transfer and subsequent environmental retention. Here, we investigated how different biocides affect the release of eDNA from mature biofilms of two opportunistic model strains Pseudomonas aeruginosa ATCC 27853 (PA) and Staphylococcus aureus ATCC 25923 (SA) and contribute to the hospital resistome in the form of surface and water contaminants and dust particles. The effect of four groups of biocides, alcohols, hydrogen peroxide, quaternary ammonium compounds, and the polymeric biocide polyhexamethylene guanidine hydrochloride (PHMG-Cl), was evaluated using PA and SA biofilms. Most biocides, except for PHMG-Cl and 70% ethanol, caused substantial eDNA release, and PHMG-Cl was found to block biofilm development when used at concentrations of 0.5% and 0.1%. This might be associated with the formation of DNA–PHMG-Cl complexes as PHMG-Cl is predicted to bind to AT base pairs by molecular docking assays. PHMG-Cl was found to bind high-molecular DNA and plasmid DNA and continued to inactivate DNA on surfaces even after 4 weeks. PHMG-Cl also effectively inactivated biofilm-associated antibiotic resistance gene eDNA released by a pan-drug-resistant Klebsiella strain, which demonstrates the potential of a polymeric biocide as a new surface-active agent to combat the spread of antibiotic resistance in hospital settings.

Novel antibiotic combinations may act synergistically to inhibit the growth of multidrug-resistant bacterial pathogens but predicting which combination will be successful is difficult, and standard antimicrobial susceptibility testing may not identify important physiological differences between planktonic free-swimming and biofilm-protected surface-attached sessile cells. Using a nominally macrolide-resistant model Klebsiella pneumoniae strain (ATCC 10031) we demonstrate the effectiveness of several macrolides in inhibiting biofilm growth in multi-well plates, and the ability of azithromycin (AZM) to improve the effectiveness of the antibacterial last-agent-of-choice for K. pneumoniae infections, colistin methanesulfonate (CMS), against biofilms. This synergistic action was also seen in biofilm tests of several K. pneumoniae hospital isolates and could also be identified in polymyxin B disc-diffusion assays on azithromycin plates. Our work highlights the complexity of antimicrobial-resistance in bacterial pathogens and the need to test antibiotics with biofilm models where potential synergies might provide new therapeutic opportunities not seen in liquid culture or colony-based assays.

A therapeutic combination of azithromycin (AZM) and colistin methanesulfonate (CMS) was shown to be effective against both non-PDR and PDR Klebsiella pneumoniae biofilms in vitro. These anti-biofilm effects, however, may not correlate with effects observed in standard plate assays, nor will they representative of in vivo therapeutic action. After all, biofilm-associated infection processes are also impacted by the presence of wound bed components, such as host cells or wound fluids, which can all affect the antibiotic effectiveness. Therefore, an in vitro wound model of biofilm infection which partially mimics the complex microenvironment of infected wounds was developed to investigate the therapeutic potential of an AZM-CMS combination against XDR K. pneumoniae isolates. The model consists of a 3D collagen sponge-like scaffold seeded with HEK293 cells submerged in a fluid milieu mimicking the wound bed exudate. Media that were tested were all based on different strengths of Dulbecco’s modified Eagles/high glucose medium supplemented with fetal bovine serum, and/or Bacto Proteose peptone. Use of this model confirmed AZM to be a highly effective antibiofilm component, when applied alone or in combination with CMS, whereas CMS alone had little antibacterial effectiveness or even stimulated biofilm development. The wound model proposed here proves therefore, to be an effective aid in the study of drug combinations under realistic conditions.

Triggering receptor expressed on myeloid cells 2 (TREM2)-expressing macrophages and systemic levels of soluble TREM2 (sTREM2) appear critical in the development of chronic liver disease (CLD) and seem relevant in its detection. The aim of this study was to examine sTREM2 as a marker for early CLD and its potential to predict posthepatectomy liver failure (PHLF) in patients undergoing partial hepatectomy. sTREM2 was assessed in the plasma of 108 patients undergoing liver resection. Blood was drawn prior to surgery (preop) and on the first and fifth postoperative day. Preop sTREM2 levels were similar across different indications for resection (p = 0.091). Higher preop sTREM2 levels were associated with advanced hepatic fibrosis (p = 0.030) and PHLF (p = 0.007). Fibrosis-4 index (FIB-4) (p = 0.619) and model for end-stage liver disease (MELD) (p = 0.590) did not show a difference between patients grouped by their CLD. Comparing the AUC from receiver-operating characteristic analysis, sTREM2 (AUC = 0.708) outperformed FIB-4 (AUC = 0.529), MELD (AUC = 0.587), Child-Pugh grading (AUC = 0.570) and LiMAx (liver maximum capacity test) (AUC = 0.516) in predicting PHLF. Similarly, in uni- and multivariate analysis, only sTREM2 proved predictive for PHLF (p = 0.023). High-risk (p = 0.003) and low-risk (p = 0.011) cut-offs for systemic sTREM2 levels could identify patients at risk for adverse outcomes after surgery. Finally, high sTREM2 was associated with decreased overall survival after liver surgery (p <0.001). Circulating sTREM2 shows sensitivity for early-stage, asymptomatic liver disease, irrespective of the underlying indication for liver surgery. Assessment of CLD via sTREM2 monitoring could improve early detection of CLD and improve outcomes after liver surgery. Soluble TREM2 (sTREM2) has previously been shown to correlate with the degree of chronic liver disease. We found that even in patients undergoing liver resection, who generally do not suffer from end-stage liver disease, sTREM2 reflects liver fibrosis status and predicts postoperative development of liver dysfunction. This is especially relevant for liver surgeons and patients, as postoperative liver dysfunction is the main reason for postoperative mortality. Our findings are also important for hepatologists, as early detection of liver fibrosis and cirrhosis is paramount for overall patient survival and we can show that even in a cohort with a median model for end-stage liver disease score of 6, sTREM2 is able to distinguish patients based on their liver fibrosis status.

The choice of effective biocides used for routine hospital practice should consider the role of disinfectants in the maintenance and development of local resistome and how they might affect antibiotic resistance gene transfer within the hospital microbial population. Currently, there is little understanding of how different biocides contribute to eDNA release that may contribute to gene transfer and subsequent environmental retention. Here we investigated how different biocides affected the release of eDNA from mature biofilms of two opportunistic model strains Pseudomonas aeruginosa ATCC 27853 (PA) and Staphylococcus aureus ATCC 25923 (SA) and contribute to the hospital resistome in the form of surface and water contaminants and dust particles. The effect of four groups of biocides including alcohols, hydrogen peroxide, quaternary ammonium compounds, and polymeric guanidines were evaluated using PA and SA biofilms. Most biocides, except for PHMG-Cl and 70% ethanol, caused substantial eDNA release and PHMG-Cl was found to block biofilm development when used at concentrations of 0.5% and 0.1%. This might be associated with the formation of DNA-PHMG-Cl complexes as PHMG-Cl is predicted to bind to AT base pairs by molecular docking assays. PHMG-Cl was found to bind high molecular DNA and plasmid DNA and continued to inactivate DNA on surfaces even after four weeks. PHMG-Cl also effectively inactivated biofilm-associated antibiotic resistance gene eDNA released by a pan-drug-resistant Klebsiella strain which demonstrates the potential of PHMG-Cl as a new surface-active agent to combat the spread of antibiotic resistance in hospital settings.

Recent reclassification of steatotic liver disease (SLD) distinguishes metabolic dysfunction-associated steatotic liver disease (MASLD) from MetALD, a newly defined entity combining MASLD with alcohol consumption. Since the mechanisms linking alcohol consumption in the context of SLD to cardiovascular disease (CVD), the leading cause of SLD mortality, remain elusive, we investigated how metabolic dysregulation and alcohol intake synergistically promote atherosclerosis. Low-density lipoprotein receptor-deficient (Ldlr^-/-) mice were fed a high-fat, high-cholesterol (HFC) diet with regular or ethanol-containing drinking water (10-20% v/v). Germ-free and antibiotic-treated Ldlr^-/- mice were used to assess the contribution of ethanol-induced dysbiosis. Associations between alcohol consumption and cardiometabolic risk were assessed in two human cohorts (N = 5115, N = 2515). Ethanol intake in HFC diet-fed Ldlr^-/- mice exacerbated hepatic steatosis and systemic dyslipidemia, despite only modest elevations in systemic ethanol levels (ps≤0.05). Liver transcriptomic profiling revealed ethanol-induced alterations in lipid metabolism and enhanced proinflammatory signatures, accompanied by increased recruitment of Ly6C^high monocytes to the liver (p=0.0121) and elevated levels in circulation (p=0.0043). Correspondingly, ethanol-consuming HFC diet-fed Ldlr^-/- mice developed enlarged aortic root lesions (p=0.0105). Neither germ-free conditions nor antibiotic treatment mitigated CVD progression. Metabolomic profiling revealed hyperuricemia in ethanol-exposed HFC diet-fed Ldlr^-/- mice, which was associated with an upregulation of inflammasome-related genes in the liver, along with an increase in hepatic NLRP3 protein expression (ps≤0.05). Notably, human data mirrored these findings, demonstrating a dose-dependent association between alcohol intake, dyslipidemia, monocytosis, hyperuricemia, and increased cardiovascular risk (ps≤0.05). Our findings identify alcohol as an important immune-modulatory lifestyle factor that contributes to elevated cardiovascular risk. Alcohol consumption, even at moderate levels, contributes to cardiovascular risk, particularly in individuals with SLD. Combining murine models with human cohort data, we show that alcohol intake in SLD is associated with coordinated immune-metabolic alterations, including dyslipidemia, monocytosis, hyperuricemia, and enhanced NLRP3 inflammasome signaling, collectively indicating an elevated cardiovascular risk profile. These findings are relevant to hepatology and cardiovascular medicine, linking alcohol consumption to measurable systemic pathways beyond liver injury. Clinically, systematic assessment of alcohol intake and incorporation into cardiovascular risk evaluation may improve risk stratification and support closer surveillance and preventive strategies in patients with SLD.