The biofilm-forming microbial species Candida parapsilosis and Staphylococcus epidermidis have been recently linked to serious infections associated with implanted medical devices. We studied microbial biofilms by high resolution scanning electron microscopy (SEM), which allowed us to visualize the biofilm structure, including the distribution of cells inside the extracellular matrix and the areas of surface adhesion. We compared classical SEM (chemically fixed samples) with cryogenic SEM, which employs physical sample preparation based on plunging the sample into various liquid cryogens, as well as high-pressure freezing (HPF). For imaging the biofilm interior, we applied the freeze-fracture technique. In this study, we show that the different means of sample preparation have a fundamental influence on the observed biofilm structure. We complemented the SEM observations with Raman spectroscopic analysis, which allowed us to assess the time-dependent chemical composition changes of the biofilm in vivo. We identified the individual spectral peaks of the biomolecules present in the biofilm and we employed principal component analysis (PCA) to follow the temporal development of the chemical composition.

• Keywords
• Raman spectroscopy, biofilm, cryo-SEM, sample preparation, scanning electron microscopy,
• MeSH
• Bacterial Infections diagnosis   microbiology   MeSH
• Biofilms growth & development   MeSH
• Candida parapsilosis isolation & purification   pathogenicity   ultrastructure   MeSH
• Humans MeSH
• Microscopy, Electron, Scanning MeSH
• Spectrum Analysis, Raman MeSH
• Staphylococcus epidermidis isolation & purification   pathogenicity   ultrastructure   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH