Establishment of an in vitro bacterial/fungal biofilm model composed of Streptococcus pyogenes and Candida albicans.

• Grant number: 22/09156-2
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: October 01, 2022
• End date: September 30, 2023
• Field of knowledge: Health Sciences - Dentistry - Dental Clinics
• Principal Investigator: Douglas Roberto Monteiro
• Grantee: Amanda Munarolo Piacenza de Oliveira
• Host Institution: Faculdade de Ciências da Saúde (FCSA). Universidade do Oeste Paulista (UNOESTE). Presidente Prudente , SP, Brazil

Abstract

The aim of the present study is to establish an in vitro model of a mixed biofilm of Streptococcus pyogenes and Candida albicans, determining the ideal conditions for its formation. Mixed biofilms will be formed for 48 hours on glass discs vertically positioned in the Amsterdam Active Attachment Model. Different factors of variation will be evaluated, including the culture medium (Brain Heart Infusion broth or artificial saliva), the human salivary pellicle (present or absent) on the glass discs prior to biofilm formation, and the sequence of inoculation of the microorganisms (inoculation of S. pyogenes followed by C. albicans, inoculation of C. albicans followed by S. pyogenes or inoculation of both species at the same time), generating 12 different models of biofilm formation. The resulting biofilms will be analyzed by counting colony-forming units (using specific culture media) and quantifying the total biomass (using the crystal violet staining assay) and metabolic activity (XTT reduction assay). Scanning electron microscopy and confocal laser scanning microscopy will be used to analyze the ultrastructure of the resulting biofilms. In addition, single biofilms of the two species will be tested as controls. All microbiological tests will be performed in triplicate on three different occasions. Data will be analyzed to employ the most appropriate statistical test. P < .05 will be considered statistically significant.