Cell wall thickening is a common feature among daptomycin-resistant Staphylococcus aureus strains. However, the mechanism(s) leading to this phenotype is unknown. We examined a number of cell wall synthesis pathway parameters in an isogenic strain set of S. aureus bloodstream isolates obtained from a patient with recalcitrant endocarditis who failed daptomycin therapy, including the initial daptomycin-susceptible parental strain (strain 616) and two daptomycin-resistant strains (strains 701 and 703) isolated during daptomycin therapy. Transmission electron microscopy demonstrated significantly thicker cell walls in the daptomycinresistant strains than in the daptomycin-susceptible strain, a finding which was compatible with significant differences in dry cell weight of strain 616 versus strains 701 to 703 (P < 0.05). Results of detailed analysis of cell wall muropeptide composition, the degree of peptide side chain cross-linkage, and the amount of the peptidoglycan precursor, UDP-MurNAc-pentapeptide, were similar in the daptomycin-susceptible and daptomycin-resistant isolates. In contrast, the daptomycin-resistant strains contained less O-acetylated peptidoglycan. Importantly, both daptomycin-resistant strains synthesized significantly more wall teichoic acid (WTA) than the parental strain (P < 0.001). Moreover, the proportion of D-alanylated WTA species was substantially higher in the daptomycin-resistant strains than in the daptomycin-susceptible parental strain (P < 0.05 in comparing strain 616 versus strain 701). The latter phenotypic findings correlated with (i) enhanced tagA and dltA gene expression, respectively, and (ii) an increase in surface positive charge observed in the daptomycinresistant versus daptomycin-susceptible isolates. Collectively, these data suggest that increases in WTA synthesis and the degree of its D-alanylation may play a major role in the daptomycin-resistant phenotype in some S. aureus strains.Daptomycin has become a key agent for the management of serious Staphylococcus aureus infections, especially for drug-resistant strains, such as methicillin-resistant S. aureus (MRSA) (21, 35). However, a number of recent reports have documented the emergence of in vivo daptomycin-resistant S. aureus strains during unsuccessful therapy with this agent (3,11,27). There appear to be several potential mechanisms associated with the daptomycin-resistant phenotype at both the genotypic and phenotypic levels, including (i) increased expression of genes involved in maintenance of the bacterial surface positive charge (e.g., dltABCD or mprF [37,38,40]), (ii) perturbations in cell membrane fluidity (18), and (iii) altered cell membrane permeabilization (12). We have recently shown that a common (although not universal) accompaniment of the daptomycin-resistant phenotype is a notably thickened cell wall among such strains (18,37,38). Although some investigations have shown the presence of selected cell wall synthetic alterations in daptomycin-resistant S. aureus strains (e.g., changes in peptidoglycan...

Nasal colonization is a major risk factor for S. aureus infections. The mechanisms responsible for colonization are still not well understood and involve several factors on the host and the bacterial side. One key factor is the cell wall teichoic acid (WTA) of S. aureus, which governs direct interactions with nasal epithelial surfaces. We report here the first receptor for the cell wall glycopolymer WTA on nasal epithelial cells. In several assay systems this type F-scavenger receptor, termed SREC-I, bound WTA in a charge dependent manner and mediated adhesion to nasal epithelial cells in vitro. The impact of WTA and SREC-I interaction on epithelial adhesion was especially pronounced under shear stress, which resembles the conditions found in the nasal cavity. Most importantly, we demonstrate here a key role of the WTA-receptor interaction in a cotton rat model of nasal colonization. When we inhibited WTA mediated adhesion with a SREC-I antibody, nasal colonization in the animal model was strongly reduced at the early onset of colonization. More importantly, colonization stayed low over an extended period of 6 days. Therefore we propose targeting of this glycopolymer-receptor interaction as a novel strategy to prevent or control S. aureus nasal colonization.

Streptococcus pneumoniae has unusually complex cell wall teichoic acid and lipoteichoic acid, both of which contain a ribitol phosphate moiety. The lic region of the pneumococcal genome contains genes for the uptake and activation of choline, the attachment of phosphorylcholine to teichoic acid precursors, and the transport of these precursors across the cytoplasmic membrane. The role of two other, so far uncharacterized, genes, spr1148 and spr1149, in the lic region was determined. TarJ (spr1148) encodes an NADPH-dependent alcohol dehydrogenase for the synthesis of ribitol 5-phosphate from ribulose 5-phosphate. TarI (spr1149) encodes a cytidylyl transferase for the synthesis of cytidine 5-diphosphate (CDP)-ribitol from ribitol 5-phosphate and cytidine 5-triphosphate. We also present the crystal structure of TarI with and without bound CDP, and the structures present a rationale for the substrate specificity of this key enzyme. No transformants were obtained with insertion plasmids designed to interrupt the tarIJ genes, indicating that their function could be essential for cell growth. CDP-activated ribitol is a precursor for the synthesis of pneumococcal teichoic acids and some of the capsular polysaccharides. Thus, all eight genes in the lic region have a role in teichoic acid synthesis.Teichoic acids are the major cell wall components of most gram-positive bacteria (36); they are made up of anionic polymers of glycerol phosphate or ribitol phosphate, with esterlinked D-alanine or sugar additions. There are two types of teichoic acids: wall teichoic acid (WTA), which is covalently linked via a phosphodiester bond to C-6 of N-acetylmuramic acid in the peptidoglycan layer, and lipoteichoic acid (LTA), which contains a terminal glycolipid that acts as an anchor within the cytoplasmic membrane. A number of important physiological functions have been assigned to teichoic acids, including cation homeostasis; trafficking of ions, nutrients, proteins, and antibiotics; regulation of autolysins; and binding of envelope proteins (36,50). LTA has recently been shown to be essential for cell growth in Staphylococcus aureus (20). Teichoic acids and their structural modifications also play important roles in the interaction of pathogenic bacteria with host organisms. For example, an S. aureus mutant lacking WTA, although showing normal growth in the laboratory, has strongly reduced capability for nasal colonization in a cotton rat model (49). Furthermore, a Streptococcus pneumoniae mutant lacking the choline modification in WTA and LTA shows drastically reduced virulence in different animal models of infection (27).The WTA and LTA of the human pathogen S. pneumoniae (the pneumococcus) are unique among bacterial teichoic acids with respect to several features. First, unlike in most other species, the repeating units of WTA and LTA in the pneumococci have identical chemical structures (18). Second, these repeating units contain the amino alcohol choline, which has been detected rarely in bacteria and is an essential growth fac...

Three new members of the fluostatin family, fluostatins CϳE, were discovered in a culture filtrate extract of strain Acta 1383 during an HPLC screening program. The producing strain belongs to the genus Streptomyces and is closely related to type strains classified in the Streptomyces lavendulae 16S rRNA subclade. Fluostatins are named by their characteristic fluorenone chromophore. Fluostatin C shows moderate activity against selected human tumor cell lines.