Uncovering Roles of Streptococcus gordonii SrtA-Processed Proteins in the Biofilm Lifestyle

Abstract

<i>Streptococcus gordonii</i> is a commensal oral organism. Harmless in the oral cavity, <i>S. gordonii</i> is an opportunistic pathogen. <i>S. gordonii</i> adheres to body surfaces using surface adhesive proteins (adhesins), which are critical to subsequent formation of biofilm communities. As in most Gram-positive bacteria, <i>S. gordonii</i> surface proteins containing the C-terminal LPXTG motif cleavage sequence are processed by sortase A (SrtA) to become covalently attached to the cell wall. To characterize the functional diversity and redundancy in the family of SrtA-processed proteins, an <i>S. gordonii</i> DL1 markerless deletion mutant library was constructed of each of the 26 putative SrtA-processed proteins. Each library member was evaluated for growth in rich medium, biofilm formation on plastic, saliva and salivary fractions, cell surface hydrophobicity (CSH), hemagglutination, and integration into an <i>ex vivo</i> plaque biofilm community. Library members were compared to the non-SrtA-processed adhesins AbpA and AbpB. While no major growth differences in rich medium were observed, many <i>S. gordonii</i> LPXTG/A proteins impacted biofilm formation on one or more of the substrates. Several mutants showed significant differences in hemagglutination, hydrophobicity, or fitness in the <i>ex vivo</i> plaque model. From the identification of redundant and unique functions in these <i>in vitro</i> and <i>ex vivo</i> systems, functional stratification among the LPXTG/A proteins is apparent.<b>IMPORTANCE</b><i>S. gordonii</i> interactions with its environment depend on the complement of cell wall proteins. A subset of these cell wall proteins requires processing by the enzyme sortase A (SrtA). The identification of SrtA-processed proteins and their functional characterization will help the community to better understand how <i>S. gordonii</i> engages with its surroundings, including other microbes, integrates into the plaque community, adheres to the tooth surface, and hematogenously disseminates to cause blood-borne infections. This study identified 26 putative SrtA-processed proteins through creation of a markerless deletion mutant library. The library was subject to functional screens that were chosen to better understand key aspects of <i>S. gordonii</i> physiology and pathogenesis.