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Microbe entry through catheter ports can lead to biofilm accumulation and complications from catheter-related bloodstream infection and ultimately require antimicrobial treatment and catheter replacement. Although strides have been made with microbial prevention by applying standardized antiseptic techniques during catheter implantation, both bacterial and fungal microbes can present health risks to already sick individuals. To reduce microbial adhesion, murine and human catheters were coated with polyurethane and auranofin using a dip coating method and compared to non-coated materials. Upon passage of fluid through the coated material in vitro , flow dynamics were not impacted. The unique antimicrobial properties of the coating material auranofin has shown inhibitory activity against bacteria such as Staphylococcus aureus and fungi such as Candida albicans . Auranofin coating on catheters at 10mg/mL reduced C . albicans accumulation in vitro from 2.0 x 10 8 to 7.8 x 10 5 CFU for mouse catheters and from 1.6 x 10 7 to 2.8 x 10 6 for human catheters, showing an impact to mature biofilms. Assessment of a dual microbe biofilm on auranofin-coated catheters resulted in a 2-log reduction in S . aureus and a 3-log reduction in C . albicans compared to uncoated catheters. In vivo assessment in a murine subcutaneous model demonstrated that catheters coated with 10 mg/mL auranofin reduced independent S . aureus and C . albicans accumulation by 4-log and 1-log, respectively, compared to non-coated catheters. In conclusion, the auranofin-coated catheters demonstrate proficiency at inhibiting multiple pathogens by decreasing S . aureus and C . albicans biofilm accumulation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Felix, Whitely, Tharmalingam, Mishra, Vera-Gonzalez, Mylonakis, Shukla and Fuchs.)

Candida albicans is the main fungal species associated with the development of oral candidiasis. Currently, therapeutic options for these infections are limited by the adverse effects of antifungal drugs and by the emergence of drug resistant strains. Thus, the development of new antifungal agents is needed for the prevention and treatment of oral Candida infections. Caffeic acid phenethyl ester (CAPE) is a natural compound from propolis polyphenolic groups that exhibits many pharmacological properties. In this study, we investigated whether CAPE can have antifungal and immunomodulatory effects on oral candidiasis. Preliminary tests to assess the antifungal activity of CAPE were performed using the Minimum Inhibitory Concentration (MIC) assay that demonstrated inhibition in a range from 16 to 32 μg/mL, confirming its antifungal activity on several C. albicans strains isolated from the oral cavity. Subsequently, we analyzed Candida spp biofilms formed in vitro , in which CAPE treatment at 5 x MIC caused a reduction of 68.5% in the total biomass and ~2.60 Log in the viable cell count (CFU/mL) in relation to the untreated biofilm ( p <0.0001). Next, RNA was extracted from untreated and CAPE-treated biofilms and analyzed by real-time qPCR. A series of genes analyzed ( ALS1, ECE1, EPA1, HWP1 , YWP1, BCR1, BGR1, CPH1, EFG1 , NDT80, ROB1, TEC1 , UME6 , SAP2 , SAP5 , PBL2 , and LIP9 ) were downregulated by CAPE compared to the untreated control group ( p <0.0001). In in vivo studies using Galleria mellonella , the treatment with CAPE prolonged survival of larvae infected by C. albicans by 44.5% ( p < 0.05) and accompanied by a 2.07-fold increase in the number of hemocytes. Flow cytometry revealed the most prominent increases were in types P2 and P3 hemocytes, granular cells, which phagocytize pathogens. In addition, CAPE treatment decreased the fungal load in the hemolymph and stimulated the expression of antifungal peptide genes such as galiomicin and gallerimycin . The antifungal and immunomodulatory activities observed in G. mellonella were extended to a murine model of oral candidiasis, in which CAPE decreased the levels of C. albicans colonization (~2 log CFU/mL) in relation to the untreated control group. In addition, CAPE treatment significantly reduced pseudomembranous lesions, invasion of hyphae on epithelium surfaces, tissue damage and inflammatory infiltrate ( p < 0.05). CAPE was also able to increase the expression of β-defensin 3 compared to the infected and untreated group by 3.91-fold ( p < 0.0001). Taken together, these results show that CAPE has both antifungal and immunomodulatory effects, making it a promising natural antifungal agent for the treatment and prevention of candidiasis and shows impact to oral candidiasis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 de Barros, Rossoni, Garcia, Kaminski, Loures, Fuchs, Mylonakis and Junqueira.)

Candida auris has emerged as a medically important pathogen with considerable resistance to antifungal agents. The ability to produce biofilms is an important pathogenicity feature of this species that aids escape of host immune responses and antimicrobial agents. The objective of this study was to verify antifungal action using in vitro and in vivo models of the Lactobacillus paracasei 28.4 probiotic cells and postbiotic activity of crude extract (LPCE) and fraction 1 (LPF1), derived from L. paracasei 28.4 supernatant. Both live cells and cells free supernatant of L. paracasei 28.4 inhibited C. auris suggesting probiotic and postbiotic effects. The minimum inhibitory concentration (MIC) for LPCE was 15 mg/mL and ranges from 3.75 to 7.5 mg/mL for LPF1. Killing kinetics determined that after 24 h treatment with LPCE or LPF1 there was a complete reduction of viable C. auris cells compared to fluconazole, which decreased the initial inoculum by 1-logCFU during the same time period. LPCE and LPF1 significantly reduced the biomass ( p = 0.0001) and the metabolic activity ( p = 0.0001) of C. auris biofilm. There was also a total reduction (~10 8 CFU/mL) in viability of persister C. auris cells after treatment with postbiotic elements ( p < 0.0001). In an in vivo study, injection of LPCE and LPF1 into G. mellonella larvae infected with C. auris prolonged survival of these insects compared to a control group ( p < 0.05) and elicited immune responses by increasing the number of circulating hemocytes and gene expression of antimicrobial peptide galiomicin. We concluded that the L. paracasei 28.4 cells and postbiotic elements (LPCE and LPF1) have antifungal activity against planktonic cells, biofilms, and persister cells of C. auris . Postbiotic supplementation derived from L. paracasei 28.4 protected G. mellonella infected with C. auris and enhanced its immune status indicating a dual function in modulating a host immune response., (Copyright © 2020 Rossoni, de Barros, Mendonça, Medina, Silva, Fuchs, Junqueira and Mylonakis.)

Intravascular catheter related bloodstream infections (CRBSIs) are a leading cause of hospital-acquired infections worldwide, resulting not only in the burden of cost and morbidity for patients but also in the over-consumption of medical resources for hospitals and health care organizations. In this study, a novel auranofin releasing antibacterial and antibiofilm polyurethane (PU) catheter coating was developed and investigated for future use in preventing CRBSIs. Auranofin is an antirheumatic drug with recently identified antimicrobial properties. The drug carrier, PU, acts as a barrier surrounding the antibacterial agent, auranofin, to extend the drug release profile and improve its long-term antibacterial and antibiofilm efficacy and potentially the length of catheter implantation within a patient. The PU+auranofin coatings developed here were found to be highly stretchable (exhibiting ~500% percent elongation), which is important for the compliance of the material on a flexible catheter. PU+auranofin coated catheters were able to inhibit the growth of methicillin-resistant Staphylococcus aureus (MRSA) for 8 to 26 days depending on the specific drug concentration utilized during the dip coating process. The PU+auranofin coated catheters were also able to completely inhibit MRSA biofilm formation in vitro , an effect that was not observed with auranofin or PU alone. Lastly, these coatings were found to be hemocompatible with human erythrocytes and maintain liver cell viability.

Systemic Candida infections remain a leading cause of nosocomial infections in the United States and worldwide. Many challenges remain in achieving rapid, direct diagnosis of fungal bloodstream infections due to limitations of conventional diagnostic methods that continue to demonstrate poor sensitivity, prolonged culture times that lead to delayed treatment, and detection variability between tests that compromises result reproducibility. Despite advancements in technology, mortality, and cost of care presented by blood stream infection with Candida spp. (candidemia) continues to rise and there is an urgent need for the development of novel methods to accurately detect Candida species present within the blood. This is especially true when patients are infected with drug resistant strains of Candida where accurate and immediate therapeutic treatment is of the importance. This study presents a method of separating fungal cells from lysed blood using inertial forces applied through microfluidics in order to abbreviate the time required to achieve a diagnosis by mitigating the need to grow blood cultures. We found that C. albicans can segregate into a focused stream distinct from white blood cells isolated within the Inertial Fungal Focuser (IFF) after red blood cell lysis. As a result of the focusing process, the collected cells are also concentrated 2.86 times. The same IFF device is applicable to non- albicans species: Candida parapsilosis, Candida glabrata , and Candida tropicalis , providing both isolation from lysed blood and a reduction in solution volume. Thus, the devised platform provides a means to isolate medically significant fungal cells from blood and concentrate the cells for further interrogation.