Streptococcus pneumoniae, S. mitis, and S. oralis Produce a Phosphatidylglycerol-Dependent, <i>ltaS</i> -Independent Glycerophosphate-Linked Glycolipid

Abstract

Lipoteichoic acid (LTA) is a Gram-positive bacterial cell surface polymer that participates in host-microbe interactions. It was previously reported that the major human pathogen <i>Streptococcus pneumoniae</i> and the closely related oral commensals <i>S. mitis</i> and <i>S. oralis</i> produce type IV LTAs. Herein, using liquid chromatography/mass spectrometry-based lipidomic analysis, we found that in addition to type IV LTA biosynthetic precursors, <i>S. mitis</i>, <i>S. oralis</i>, and <i>S. pneumoniae</i> also produce glycerophosphate (Gro-P)-linked dihexosyl (DH)-diacylglycerol (DAG), which is a biosynthetic precursor of type I LTA. <i>cdsA</i> and <i>pgsA</i> mutants produce DHDAG but lack (Gro-P)-DHDAG, indicating that the Gro-P moiety is derived from phosphatidylglycerol (PG), whose biosynthesis requires these genes. <i>S. mitis</i>, but not <i>S. pneumoniae</i> or <i>S. oralis</i>, encodes an ortholog of the PG-dependent type I LTA synthase, <i>ltaS</i> By heterologous expression analyses, we confirmed that <i>S. mitis</i><i>ltaS</i> confers poly(Gro-P) synthesis in both <i>Escherichia coli</i> and <i>Staphylococcus aureus</i> and that <i>S. mitis</i><i>ltaS</i> can rescue the growth defect of an <i>S. aureus</i><i>ltaS</i> mutant. However, we do not detect a poly(Gro-P) polymer in <i>S. mitis</i> using an anti-type I LTA antibody. Moreover, Gro-P-linked DHDAG is still synthesized by an <i>S. mitis</i><i>ltaS</i> mutant, demonstrating that <i>S. mitis</i> LtaS does not catalyze Gro-P transfer to DHDAG. Finally, an <i>S. mitis</i><i>ltaS</i> mutant has increased sensitivity to human serum, demonstrating that <i>ltaS</i> confers a beneficial but currently undefined function in <i>S. mitis</i> Overall, our results demonstrate that <i>S. mitis</i>, <i>S. pneumoniae</i>, and <i>S. oralis</i> produce a Gro-P-linked glycolipid via a PG-dependent, <i>ltaS</i>-independent mechanism.<b>IMPORTANCE</b> The cell wall is a critical structural component of bacterial cells that confers important physiological functions. For pathogens, it is a site of host-pathogen interactions. In this work, we analyze the glycolipids synthesized by the mitis group streptococcal species, <i>S. pneumoniae</i>, <i>S. oralis</i>, and <i>S. mitis</i> We find that all produce the glycolipid, glycerophosphate (Gro-P)-linked dihexosyl (DH)-diacylglycerol (DAG), which is a precursor for the cell wall polymer type I lipoteichoic acid in other bacteria. We investigate whether the known enzyme for type I LTA synthesis, LtaS, plays a role in synthesizing this molecule in <i>S. mitis</i> Our results indicate that a novel mechanism is responsible. Our results are significant because they identify a novel feature of <i>S. pneumoniae</i>, <i>S. oralis</i>, and <i>S. mitis</i> glycolipid biology.