Human serum induces daptomycin tolerance in Enterococcus faecalis and viridans group streptococci

Daptomycin is a membrane-targeting lipopeptide antibiotic used in the treatment of infective endocarditis caused by multidrug-resistant Gram-positive bacteria such as Staphylococcus aureus, enterococci and viridans group streptococci. Despite demonstrating excellent in vitro activity and a low prevalence of resistant isolates, treatment failure is a significant concern, particularly for enterococcal infection. We have shown recently that human serum triggers daptomycin tolerance in S. aureus, but it was not clear if a similar phenotype occurred in other major infective endocarditis pathogens. We found that Enterococcus faecalis, Streptococcus gordonii or Streptococcus mutans grown under standard laboratory conditions were efficiently killed by dapto-mycin, whereas bacteria pre-incubated in human serum survived exposure to the antibiotic, with >99 % cells remaining viable. Incubation of enterococci or streptococci in serum led to peptidoglycan accumulation, as shown by increased incorporation of the fluorescent d-amino acid analogue HADA. Inhibition of peptidoglycan accumulation using the antibiotic fosfomycin resulted in a >tenfold reduction in serum-induced daptomycin tolerance, demonstrating the important contribution of the cell wall to the phenotype. We also identified a small contribution to daptomycin tolerance in E. faecalis from cardiolipin synthases, although this may reflect the inherent increased susceptibility of cardiolipin-deficient mutants. In summary, serum-induced daptomycin tolerance is a consistent phenomenon between Gram-positive infective endocarditis pathogens, but it may be mitigated using currently available antibiotic combination therapy.