Federico C. Beasley

Iron is critical for virtually all forms of life. The production of high-affinity iron chelators, siderophores, and the subsequent uptake of iron-siderophore complexes are a common strategy employed by microorganisms to acquire iron. Staphylococcus aureus produces siderophores but genetic information underlying their synthesis and transport is limited. Previous work implicated the sbn operon in siderophore synthesis and the sirABC operon in uptake. Here we characterize a second siderophore biosynthetic locus in S. aureus; the locus consists of four genes (in strain Newman these open reading frames are designated NWMN_2079-2082) which, together, are responsible for the synthesis and export of staphyloferrin A, a polycarboxylate siderophore. While deletion of the NWMN_2079-2082 locus did not affect iron-restricted growth of S. aureus, strains bearing combined sbn and NWMN_2079-2082 locus deletions produced no detectable siderophore and demonstrated severely attenuated iron-restricted growth. Adjacent to NWMN_2079-2082 resides the htsABC operon, encoding an ABC transporter previously implicated in haem acquisition. We provide evidence here that HtsABC, along with the FhuC ATPase, is required for the uptake of staphyloferrin A. The crystal structure of apo-HtsA was determined and identified a large positively charged region in the substrate-binding pocket, in agreement with a role in binding of anionic staphyloferrin A.

The tremendous success of Staphylococcus aureus as a pathogen is due to the controlled expression of a diverse array of virulence factors. The effects of host environments on the expression of virulence factors and the mechanisms by which S. aureus adapts to colonize distinct host tissues are largely unknown. Vertebrates have evolved to sequester nutrient iron from invading bacteria, and iron availability is a signal that alerts pathogenic microorganisms when they enter the hostile host environment. Consistent with this, we report here that S. aureus senses alterations in the iron status via the ferric uptake regulator (Fur) and alters the abundance of a large number of virulence factors. These Fur-mediated changes protect S. aureus against killing by neutrophils, and Fur is required for full staphylococcal virulence in a murine model of infection. A potential mechanistic explanation for the impact of Fur on virulence is provided by the observation that Fur coordinates the reciprocal expression of cytolysins and a subset of immunomodulatory proteins. More specifically, S. aureus lacking fur exhibits decreased expression of immunomodulatory proteins and increased expression of cytolysins. These findings reveal that Fur is involved in initiating a regulatory program that organizes the expression of virulence factors during the pathogenesis of S. aureus pneumonia.

Staphylococcus aureus is a frequent cause of bloodstream, respiratory tract, and skin and soft tissue infections. In the bloodstream, the iron-binding glycoprotein transferrin circulates to provide iron to cells throughout the body, but its iron-binding properties make it an important component of innate immunity. It is well established that siderophores, with their high affinity for iron, in many instances can remove iron from transferrin as a means to promote proliferation of bacterial pathogens. It is also established that catecholamine hormones can interfere with the iron-binding properties of transferrin, thus allowing infectious bacteria access to this iron pool. The present study demonstrates that S. aureus can use either of two carboxylate-type siderophores, staphyloferrin A and staphyloferrin B, via the transporters Hts and Sir, respectively, to access the transferrin iron pool. Growth of staphyloferrin-producing S. aureus in serum or in the presence of holotransferrin was not enhanced in the presence of catecholamines. However, catecholamines significantly enhanced the growth of staphyloferrin-deficient S. aureus in human serum or in the presence of human holotransferrin. It was further demonstrated that the Sst transporter was essential for this activity as well as for the utilization of bacterial catechol siderophores. The substrate binding protein SstD was shown to interact with ferrated catecholamines and catechol siderophores, with low to submicromolar affinities. Experiments involving mice challenged intravenously with wild-type S. aureus and isogenic mutants demonstrated that the combination of Hts, Sir, and Sst transport systems was required for full virulence of S. aureus.