Host Fatty Acid Utilization by Staphylococcus aureus at the Infection Site

Abstract

Staphylococcus aureus utilizes the fatty acid (FA) kinase system to activate exogenous FAs for membrane synthesis. We developed a lipidomics workflow to determine the membrane phosphatidylglycerol (PG) molecular species synthesized by S. aureus at the thigh infection site. Wild-type S. aureus utilizes both host palmitate and oleate to acylate the 1 position of PG, and the 2 position is occupied by pentadecanoic acid arising from de novo biosynthesis. Inactivation of FakB2 eliminates the ability to assimilate oleate and inactivation of FakB1 reduces the content of saturated FAs and enhances oleate utilization. Elimination of FA activation in either ΔfakA or ΔfakB1 ΔfakB2 mutants does not impact growth. All S. aureus strains recovered from the thigh have significantly reduced branched-chain FAs and increased even-chain FAs compared to that with growth in rich laboratory medium. The molecular species pattern observed in the thigh was reproduced in the laboratory by growth in isoleucine-deficient medium containing exogenous FAs. S. aureus utilizes specific host FAs for membrane biosynthesis but also requires de novo FA biosynthesis initiated by isoleucine (or leucine) to produce pentadecanoic acid. IMPORTANCE The shortage of antibiotics against drug-resistant Staphylococcus aureus has led to the development of new drugs targeting the elongation cycle of fatty acid (FA) synthesis that are progressing toward the clinic. An objection to the use of FA synthesis inhibitors is that S. aureus can utilize exogenous FAs to construct its membrane, suggesting that the bacterium would bypass these therapeutics by utilizing host FAs instead. We developed a mass spectrometry workflow to determine the composition of the S. aureus membrane at the infection site to directly address how S. aureus uses host FAs. S. aureus strains that cannot acquire host FAs are as effective in establishing an infection as the wild type, but strains that require the utilization of host FAs for growth were attenuated in the mouse thigh infection model. We find that S. aureus does utilize host FAs to construct its membrane, but host FAs do not replace the requirement for pentadecanoic acid, a branched-chain FA derived from isoleucine (or leucine) that predominantly occupies the 2 position of S. aureus phospholipids. The membrane phospholipid structure of S. aureus mutants that cannot utilize host FAs indicates the isoleucine is a scarce resource at the infection site. This reliance on the de novo synthesis of predominantly pentadecanoic acid that cannot be obtained from the host is one reason why drugs that target fatty acid synthesis are effective in treating S. aureus infections.

Lipidomic and Ultrastructural Characterization of the Cell Envelope of Staphylococcus aureus Grown in the Presence of Human Serum

“…The same group showed that exogenous fatty acids could occupy both the sn-1 and sn-2 positions of PG when cells were grown in brain heart infusion broth supplemented with C 14:0 , C 16:0 , and C 18:1 , or serum (38). This seemingly disproves the essentiality of the requirement for biosynthesized fatty acid anteiso C 15:0 at the sn-2 position (15,36,40) and undermines the viability of inhibitors of the FASII pathway as useful therapeutic agents (38). In this work, we also observed lipid species containing no C 15:0 , such as PG 32:1 (18:1/14:0), PG 34:1 (18:1/16:0 and 20:1/14:0), and PG 36:1 (18:1/18:0, 20:1/16:0, and 22:1/14:0), which supports the notion that anteiso C 15:0 is not essential.…”

Branched chain fatty acid synthesis drives tissue-specific innate immune response and infection dynamics of Staphylococcus aureus

“…Staphylococcus aureus synthesizes or scavenges essential nutrients and evades immune responses to promote persistence during infection [1][2][3][4][5]. One such nutrient is fatty acids [6][7][8][9][10][11]. Fatty acids are essential to the structure and function of bacterial phospholipids and lipoproteins and serve to maintain membrane homeostasis [7,[12][13][14].…”

“…Incorporation of eFA from the host environment can support bacterial survival in the presence of FASII inhibitors through bypass of de novo fatty acid synthesis [12,19,25,26]. In addition, studies with human serum and murine thigh infection imply a degree of UFA incorporation by S. aureus even in the absence of FASII inhibitors [8,9]. Beyond restoring viability during FASII inhibition, the functional consequences of flexibility in membrane fatty acid composition on the virulence of S. aureus is unknown.…”

To make or take: bacterial lipid homeostasis during infection

“…PG 34:3 (16:1 + 18:2) is an unusual species, not commonly found in mammalian cells (LipidMAPS database), nor in A. baumannii grown in LB media (Fig 1i) (10). Hence, its presence in A. baumannii isolated from the mouse pleural cavity is a strong indicator of A. baumannii 18:2 acquisition from the host, reminiscent of 18:1 acquisition by S. aureus in mouse thigh tissue (11).…”

“…PG 34:3 (16:1 + 18:2) is an unusual species, not commonly found in mammalian cells (LipidMAPS database), nor in A. baumannii grown in LB media ( Fig 1i ) (10). Hence, its presence in A. baumannii isolated from the mouse pleural cavity is a strong indicator of A. baumannii 18:2 acquisition from the host, reminiscent of 18:1 acquisition by S. aureus in mouse thigh tissue (11). The enhanced incorporation of 18:2 fatty acids in the A. baumannii PE and PG pools affected the total number of unsaturated fatty acids in A. baumannii from the PL, as compared to those cultured in LB, by 1.3-fold (PE) and 1.1-fold (PG) ( Fig 1k ).…”