The need for new antimicrobials to treat bacterial infections has led to the use of type II fatty acid synthesis (FASII) enzymes as front-line targets. However, recent studies suggest that FASII inhibitors may not work against the opportunist pathogen Staphylococcus aureus, as environmental fatty acids favor emergence of multi-anti-FASII resistance. As fatty acids are abundant in the host and one FASII inhibitor, triclosan, is widespread, we investigated whether fatty acid pools impact resistance in clinical and veterinary S. aureus isolates. Simple addition of fatty acids to the screening medium led to a 50% increase in triclosan resistance, as tested in 700 isolates. Moreover, nonculturable triclosan-resistant fatty acid auxotrophs, which escape detection under routine conditions, were uncovered in primary patient samples. FASII bypass in selected isolates correlated with polymorphisms in the acc and fabD loci. We conclude that fatty-acid-dependent strategies to escape FASII inhibition are common among S. aureus isolates and correlate with anti-FASII resistance and emergence of nonculturable variants.KEYWORDS antibiotic resistance, infection, persistence, nondetectable resistance, fatty acids, Staphylococcus aureus S taphylococcus aureus is a leading cause of a wide range of infections that can affect numerous host organs and cause a range of effects from mild symptoms to severe and life-threatening diseases. Acquisition of antimicrobial resistance, likely due to overuse of antibiotics, is the principal cause of S. aureus drug resistance (1). A class of new-generation antimicrobials in intensive development uses the type II fatty acid synthesis (FASII) pathway as an antibacterial target (2-5) to treat S. aureus infections (6-8). A widely used biocide, triclosan (5-chloro-2-[2,4-dichlorophenoxy]phenol; commercialized as Irgasan or Microban), is a prototype for further anti-FASII development (4, 9, 10).The utility of FASII inhibitors was questioned when several Gram-positive bacteria were shown to be refractory to FASII inhibitors in the presence of exogenous fatty acids, making FASII enzymes dispensable (11,12). Both free and complexed fatty acids are abundant in the host (13,14), which would facilitate FASII bypass. Reservoirs and invasion sites of clinical and community-acquired staphylococci (i.e., skin, nares, gut, blood, and organs) are naturally rich in fatty acids (13-16), and triclosan is present in the environment and in human body fluids (17,18). This combination could favor FASII bypass via emergence of triclosan-resistant variants, including fatty acid auxotrophs. Fatty-acid-dependent isolates escape detection on standard isolation media, thereby confounding diagnosis and treatment.

The activity of amino acid-dependent acid resistance systems allows Escherichia coli to survive during prolonged incubation under phosphate (P i ) starvation conditions. We show in this work that rpoS-null mutants incubated in the absence of any amino acid survived during prolonged incubation under aerobic, P i starvation conditions. Whereas rpoS ؉ cells incubated with glutamate excreted high levels of acetate, rpoS mutants grew on acetic acid. The characteristic metabolism of rpoS mutants required the activity of Fur (ferric uptake regulator) in order to decrease the synthesis of the small RNA RyhB that might otherwise inhibit the synthesis of iron-rich proteins. We propose that RpoS ( S ) and the small RNA RyhB contribute to decrease the synthesis of iron-rich proteins required for the activity of the tricarboxylic acid (TCA) cycle, which redirects the metabolic flux toward the production of acetic acid at the onset of stationary phase in rpoS ؉ cells. In contrast, Fur activity, which represses ryhB, and the lack of RpoS activity allow a substantial activity of the TCA cycle to continue in stationary phase in rpoS mutants, which decreases the production of acetic acid and, eventually, allows growth on acetic acid and P i excreted into the medium. These data may help explain the fact that a high frequency of E. coli rpoS mutants is found in nature.