Endovascular Infections Caused by Methicillin-Resistant Staphylococcus aureus Are Linked to Clonal Complex-Specific Alterations in Binding and Invasion Domains of Fibronectin-Binding Protein A as Well as the Occurrence of fnbB

Abstract

e Endovascular infections caused by Staphylococcus aureus involve interactions with fibronectin present as extracellular matrix or surface ligand on host cells. We examined the expression, structure, and binding activity of the two major S. aureus fibronectinbinding proteins (FnBPA, FnBPB) in 10 distinct, methicillin-resistant clinical isolates from patients with either persistent or resolving bacteremia. The persistent bacteremia isolates (n ‫؍‬ 5) formed significantly stronger bonds with immobilized fibronectin as determined by dynamic binding measurements performed with atomic force microscopy. Several notable differences were also observed when the results were grouped by clonal complex 5 (CC5) strains (n ‫؍‬ 5) versus CC45 strains (n ‫؍‬ 5). Fibronectinbinding receptors on CC5 formed stronger bonds with immobilized fibronectin (P < 0.001). The fnbA gene was expressed at higher levels in CC45, whereas fnbB was found in only CC5 isolates. The fnbB gene was not sequenced because all CC45 isolates lacked this gene. Instead, comparisons were made for fnbA, which was present in all 10 isolates. Sequencing of fnbA revealed discrete differences within high-affinity, fibronectin-binding repeats (FnBRs) of FnBPA that included (i) 5-amino-acid polymorphisms in FnBR-9, FnBR-10, and FnBR-11 involving charged or polar side chains, (ii) an extra, 38-amino-acid repeat inserted between FnBR-9 and FnBR-10 exclusively seen in CC45 isolates, and (iii) CC5 isolates had the SVDFEED epitope in FnBR-11 (a sequence shown to be essential for fibronectin binding), while this sequence was replaced in all CC45 isolates with GIDFVED (a motif known to favor host cell invasion at the cost of reduced fibronectin binding). These complementary sequence and binding data suggest that differences in fnbA and fnbB, particularly polymorphisms and duplications in FnBPA, give S. aureus two distinct advantages in human endovascular infections: (i) FnBPs similar to that of CC5 enhance ligand binding and foster initiation of disease, and (ii) CC45-like FnBPs promote cell invasion, a key attribute in persistent endovascular infections.

Linezolid and Rifampicin Combination to Combat cfr-Positive Multidrug-Resistant MRSA in Murine Models of Bacteremia and Skin and Skin Structure Infection

“…Consistent with this, increased FN binding of cfr-positive MRSA correlated with worse outcomes of linezolid-based mono-and combination therapies in the murine bacteremia model. This observation is also in line with previous reports that the development of a hyper-adhesive FN binding phenotype contributed to persistent MRSA bacteremia and infective endocarditis (Xiong et al, 2009(Xiong et al, , 2015. In addition, we found that cfr-positive MRSA isolates adhered better than cfr-negative isolates to human endothelial cells, and this may facilitate MRSA colonization and provide an advantage in the pathogenesis of invasive MRSA infections.…”

Amino acid polymorphisms in the fibronectin-binding repeats of fibronectin-binding protein A affect bond strength and fibronectin conformation

“…Recently, we discovered nonsynonymous SNPs in high-affinity repeats of FnBPA, such as FnBR-9, that were associated with several distinct S. aureus syndromes in humans: H782Q/ K786N with cardiac device infections (12,13) and H782Q/ K786I with persistent bacteremia and infective endocarditis (14). This correlation has been externally corroborated (15).…”

“…H782Q/K786I exhibited higher affinity for immobilized Fn at fast loading rates versus the alanine peptide (⌬⌬G 0.81 kcal/ mol; Table 2), but binding was weaker than for the wild-type and Gln/Asn peptides. This is not entirely unexpected because the Gln/Ile polymorphisms (unlike the Gln/Asn polymorphisms) are associated with persistent bacteremia in patients who do not necessarily have a cardiovascular implant (14). The Gln/Ile isolates are also associated with a host invasion motif (GIDFVED) in FnBR-11 and an extra repeat of FnBR-9 giving these strains a total of 12 rather than 11 FnBRs (14,15).…”

“…These experiments were performed on the wild-type peptide and each of the double mutant peptides. We focused on the double rather than single mutants because multiple polymorphisms have been linked to disease in humans (12,14,15,47). Supplemental Table S2 shows these k off values for NTD-Fn.…”

“…Six of the 11 repeats in FnBPA exhibit high affinity toward Fn (10,11). Some of these high-affinity repeats, such as the FnBR-9 of fnbA, have been found to contain nonsynonymous single-nucleotide polymorphisms (SNPs) associated with infective endocarditis and cardiovascular device infections (12)(13)(14)(15).…”