Biofilm Formation and Phospholipase and Proteinase Production in Cryptococcus neoformans Clinical Isolates and Susceptibility towards Some Bioactive Natural Products

Abstract

Background. Cryptococcosis is one of the most common fungal infections in immunocompromised patients, which is caused by Cryptococcus neoformans. However, relatively little is known about the virulence factors of C. neoformans and the incidence of antifungal drug resistance in C. neoformans is rapidly increasing. This study was undertaken to investigate the virulence factors in C. neoformans, thymol, curcumin, piperine, gallic acid, eugenol, and plumbagin for their potential antimicrobial activity against C. neoformans. Methods. The production of phospholipase and proteinase was detected using standard methods. Biofilm formation was determined using the microtiter plate method. The broth microdilution method was used to determine the antifungal activity. The antibiofilm activity was assessed using the safranin staining method. Results. All isolates of C. neoformans produced biofilms with optical density values ranging from 0.16 to 0.89. A majority of C. neoformans isolates that were tested exhibited strong phospholipase (7/8) and proteinase (5/8) production. Plumbagin (with minimum inhibitory concentration values ranging from 4 to 16 μg/mL) showed the highest antifungal activity followed by thymol (with minimum biofilm inhibitory concentration values ranging from 8 to 64 μg/mL). In addition, plumbagin showed the highest antibiofilm activity with minimum biofilm inhibitory concentration and minimum biofilm eradication concentration values ranging from 4 to 16 μg/mL and 32 to 256 μg/mL, respectively. Conclusion. Plumbagin, compared to other natural products studied, was the most efficient in terms of antifungal and antibiofilm activities. Hence, plumbagin could be used in combination with antifungals for the development of new anticryptococcal drugs.

Antibiofilm Activity of Curcumin and Piperine and Their Synergistic Effects with Antifungals against Candida albicans Clinical Isolates

“…The minimum inhibitory concentrations (MICs) of the antifungals and natural products were determined by the method previously described [ 8 ]. The natural substances and antifungals were prepared at 4096 μ g/mL and 512 μ g/mL, respectively, and serially diluted twice with SDB in a 96-well microplate to obtain a final volume of 100 μ L. The concentrations of natural substances and antifungals ranged, respectively, from 2048 to 1 μ g/mL and 256 to 0.125 μ g/mL.…”

“…The antifungal activity of natural products was considered as follows: most active (MIC value ≤1 μ g/mL), significant activity (1 ≤ MIC value ≤10 μ g/mL), moderate (10 ≤ MIC value ≤100 μ g/mL), and inactive (100 < MIC value ≤1000 μ g/mL) [ 9 ]. The cut-off values of antifungals previously described were used for Candida albicans [ 8 ]. For fluconazole, yeast with a MIC value ≤8 μ g/mL was considered susceptible, while yeast with 32 ≥ MIC value ≥32 μ g/mL was considered as intermediate, and yeast with a MIC value ≥64 μ g/mL was considered as resistant.…”

Plumbagin inhibits fungal growth, HMGB1/LOX-1 pathway and inflammatory factors in A. fumigatus keratitis

“…Furthermore, PL displays robust and extensive antifungal activity against numerous fungi, including Fusarium graminearum and Candida albicans . It not only disrupts the cell and mitochondrial membranes of the fungi, as evidenced by a concentration-dependent increase in relative electrical conductivity but also demonstrates significant anti-biofilm activity through down-regulation of FAS1 and FAS2 expression (Hassan et al, 2016 ; Shang et al, 2019 ; Qian et al, 2022 ; Bisso et al, 2023 ; Wang et al, 2023 ). These findings indicate that PL may also possess substantial antifungal effects in A. fumigatus keratitis.…”

Biofilm Formation, Pyocyanin Production, and Antibiotic Resistance Profile of Pseudomonas aeruginosa Isolates from Wounds

“…Biofilm is defined as an assemblage of microbial cells that are attached to a surface and enclosed in an extracellular matrix principally made of polysaccharide material [ 15 – 17 ]. Biofilm is considered an important virulence factor and plays a significant role in antibiotic resistance [ 18 ].…”