A preliminary study: firstly optimisation of the in vitro biofilm formation method in Trichosporon asahii strains isolated from patients in TÜrkiye

Trichosporon asahii, a fungus found in nature and human microbiota, has recently emerged as the most common cause of disseminated and deep-seated trichosporonosis, particularly in immunocompromised patients. Biofilm formation, one of the important virulence factors of T. asahii, facilitates its colonisation and proliferation, providing the fungus with antifungal and environmental stress’s resistance. In this study, we aimed to develop a reproducible T. asahii biofilm model for our future research by optimising the inoculum concentration, adhesion-time, biofilm maturation-time, static and shaking parameters. Materials and methods. We included two clinical isolates obtained from urine samples and a reference strain in the study. For each strain, we applied 36 different experimental parameters, including, 10⁵, 10⁶ and 10⁷ CFU/mL inoculum consentrations, 60-, 90-, and 120-minutes adhesion-times, 48- and 72-hours maturation-times, and static versus shaking. We determined the biofilm formation of fungus using the most commonly used crystal violet method. We determined standard deviation values with three replicates within each experiment, and we compared all the graphs obtained by repeating these experiments twice. Results. After applying 36 different experimental parameter combinations, we determined the most optimised and reproducible parameters as 10⁷ CFU/mL inoculum concentration, 120-minute adhesion-time, 72-hour maturation-time and static. Conclusion. In optimisation studies conducted worldwide on this subject, it has been reported that biofilms formation varies in terms of quantity and duration across different countries, but there isn’t any optimisation study in Turkey. We believe that the optimisation conditions we have firstly identified in our country will shed light on our future research and other studies to be conducted in the future.