Introduction and Objectives
Staphylococcus aureus and coagulase-negative staphylococci comprises approximately 65% of infections associated with medical devices and are well known for their biofilm formatting ability. Currently, there is no efficient method for early biofilm detection. Our goal was to construct a reference map of planktonic and biofilm associated proteins of S. aureus.
S. aureus reference strain (ATCC 25923) was used to grow 24 hour planktonic, 3 day, and 12 day hydrated biofilms. Bacteria were grown in tryptic soy broth liquid medium. The Centres for Disease Control biofilm reactor was used to grow 3 day, and 12 day hydrated biofilms. Extraction and fractionation were performed using lysis buffer, ultra-membrane centrifugation, followed by reduction, alkylation and digestion steps prior to Multiplex labelling using Tandem Mass Tag (TMT) 10-plex reagent, respectively. TMT-based Mass spectrometry (MS) was performed and MS data was collected on an Orbitrap Elite Mass Spectrometer. Protein identification and relative quantitation of protein levels were performed using Proteome Discoverer (version 1.3). Biostatistical analysis was performed using the TMTPrePro R package.
Results and Discussion
We identified 1636 total secreted supernatant proteins, of which 350 and 137 proteins of 3DWB and 12DWB showed significant abundance variation from planktonic preparation, respectively. Of these, we found significantly up-regulated 59 and 71 unique proteins in this biofilm producer in between 3DWB and 12DWB, respectively. On the contrary, we found significantly down-regulated 74 and 48 unique proteins in this biofilm producer in between 3DWB and 12DWB, respectively. Therefore, a comprehensive knowledge of planktonic and biofilm associated proteins identified by S. aureus will provide a basis for future studies on the development of vaccines and diagnostic biomarkers.
In this study, we constructed an initial reference map of planktonic and various growth phase of biofilm associated proteins which might be helpful to diagnose biofilm associated infections.