DEVELOPMENT OF METHODS FOR Candida albicans BIOFILM FORMATION IN SOFT LINING MATERIAL

The literature presents several methods for Candida albicans biofilm formation in soft lining material, but there is still no consensus as to which method is the most effective. The pre-treatment in artificial saliva prior to the specimens’ contamination has been evaluated with the hypothesis that the pellicle formed on the material’s surface could benefit the adhesion of C. albicans. Another variable tested is the specimens’ position during the biofilm formation, assuming gravity could improve the adhesion of those placed in the horizontal position. In addition, other circumstance that might influence the biofilm formation is the specimens’ incubation, taking into account that maintaining the specimens in an orbital incubator could impair the adhesion of C. albicans in the initial phase. The current study aimed to describe methods to optimize the contamination of soft lining material with C. albicans biofilm. Specimens (10x1mm; n=6) of a tissue conditioner (SoftoneTM; Bosworth Co., Skokie, IL, USA) were submitted or not to a pre-treatment in artificial saliva (S) in a shaking incubator (SI) at 75 rpm and 37ºC for 2 h and were then kept in distinct positions (horizontal-H or vertical-V) in a 24-well culture plate under distinct storages (incubator-I or SI) during biofilm formation. Eight experimental groups were obtained: HI, HSI, VI, VSI; HI-S, HSI-S, VI-S, and VSI-S, without and with the pre-treatment in saliva, respectively. For cell adhesion, all specimens were immersed in a C. albicans inoculum (2x107 CFU/mL) and were kept under shaking at 75 rpm or in an incubator, both at 37°C for 90 min. Finally, the specimens were washed in PBS, immersed in RPMI-1640, and maintained at the same conditions for 48 h for biofilm formation. The degree of contamination was determined by the XTT assay at 520 nm. The absorbance data (mAbs) were submitted to 1-way ANOVA/Tukey HSD (α=0.05), once the contamination of the pre-treated specimens showed low reproducibility. The HSI group presented the highest absorbance values (p=0.000) and the VI group had the lowest cell viability (p<0.05). Intermediate values were observed in the VSI and HI groups. The best method for C. albicans biofilm formation in the tissue conditioner, which has an initial antifungal effect, was to maintain the specimens in the horizontal position in the shaking incubator throughout the experiment, without pre-treatment in artificial saliva.