Biofilms play an essential role in chronic and healthcare-associated infections and are more resistant to antimicrobials compared to their planktonic counterparts due to their (1) physiological state, (2) cell density, (3) quorum sensing abilities, (4) presence of extracellular matrix, (5) upregulation of drug efflux pumps, (6) point mutation and overexpression of resistance genes, and (7) presence of persister cells. The genes involved and their implications in antimicrobial resistance are well defined for bacterial biofilms but are understudied in fungal biofilms. Potential therapeutics for biofilm mitigation that have been reported include (1) antimicrobial photodynamic therapy, (2) antimicrobial lock therapy, (3) antimicrobial peptides, (4) electrical methods, and (5) antimicrobial coatings. These approaches exhibit promising characteristics for addressing the impending crisis of antimicrobial resistance (AMR). Recently, advances in the micro- and nanotechnology field have propelled the development of novel biomaterials and approaches to combat biofilms either independently, in combination or as antimicrobial delivery systems. In this review, we will summarize the general principles of clinically important microbial biofilm formation with a focus on fungal biofilms. We will delve into the details of some novel micro- and nanotechnology approaches that have been developed to combat biofilms and the possibility of utilizing them in a clinical setting.

The genus Candida is the most common etiological factor of opportunistic fungal infections in humans. The virulence of Candida species is due to a wide repertoire of factors, specifically, the ability to form biofilms. Medical devices such as intravenous catheters, prosthetic heart valves and surgical interventions provide pathogenic microorganisms with a surface to adhere to form biofilm. The objectives of this study were to investigate the biofilm ultrastructure of Diutina (Candida) rugosa (D. rugosa) at different developmental phases using Confocal scanning laser microscopy (CSLM) and scanning electron microscopy (SEM), quantify β-glucan, total carbohydrate and total protein in the extracellular matrix (ECM) using enzymatic β-glucan kit, phenol-sulfuric acid method and Bradford’s method, respectively, and to identify Sessile Minimum Inhibition Concentrations (SMICs) of amphotericin B, caspofungin, fluconazole, and voriconazole using serial doubling dilution. From the SEM micrographs, D. rugosa biofilms were composed of adherent yeast cells and blastospores with hyphal elements. The ultrastructure of the yeast cells was collapsed and disfigured upon exposure to amphotericin B, fluconazole and voriconazole and the biofilms presented with punctured yeast morphology upon exposure to caspofungin at their respective SMICs. The matrix thickness of embedded yeast cells from CLSM micrographs was 3.9µm at 48h. However, there was reduction in the thickness of the biofilms upon antifungal exposure. The antifungal exposed biofilms exhibit bright, diffuse, green-yellow fluorescence that were not seen in the control. D. rugosa biofilm matrices revealed 172.57µg/mL of carbohydrate, and 27.11µg/mL of protein content. The β-glucan yield in D. rugosa complex planktonic cells were in the range of 2.5 to 4.38%, on the contrary, β-glucan was not detected in the ECM. The SMICs of Diutina biofilm for amphotericin B is 1024μg/mL, caspofungin is 512 μg/mL, whereas fluconazole and voriconazole is 2048 μg/mL, respectively.

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