Resistance studies with daptomycin

Abstract

We studied the in vitro emergence of resistance to daptomycin using three methods: spontaneous resistance incidence, serial passage in the presence of increasing drug concentrations, and chemical mutagenesis. No spontaneously resistant mutants were obtained for any organism tested (<10(-10) for Staphylococcus aureus, <10(-9) for Staphylococcus epidermidis, <10(-9) for Enterococcus faecalis, <10(-9) for Enterococcus faecium, <10(-8) for Streptococcus pneumoniae). Population analysis demonstrated that bacterial susceptibility to daptomycin is heterogeneous. Assay results were sensitive to calcium concentration and culture density, both of which can affect apparent resistance rates. Stable S. aureus mutants were isolated by both serial passage in liquid media and chemical mutagenesis. The daptomycin MICs for these isolates were 8- to 32-fold higher than for the parental strain. Many mutants with high MICs (>12.5 microg/ml) had significant growth defects but did not display phenotypes typical of S. aureus small colony variants. The voltage component (Delta psi) of the bacterial membrane potential was increased in three independent resistant isolates. In vivo data showed that some daptomycin-resistant mutants had lost significant virulence. For other mutants, the degree of in vitro resistance was greater than the change in in vivo susceptibility. These results suggest that infection with some daptomycin-resistant organisms may still be easily treatable.

FIG. 1

Susceptibility to daptomycin is heterogeneous in S. aureus. (A) Susceptibility is heterogeneous in both laboratory (Sa42) and clinical (Sa675) isolates (●, Sa42; ○, Sa675). Data are mean values ± the standard deviation. (B) Heterogeneous susceptibility in Sa42 is affected by calcium (○, MHA [MIC = 1 μg/ml]; ●, MHA plus 50 mg of Ca²⁺ per liter [MIC = 0.3 μg/ml]). Data are mean values ± the standard deviation. (C) Heterogeneous susceptibility is observed in daptomycin-resistant mutants (□, Sa42 [parent]; ■, Sa278 [class 1]; ○, Sa295 [class 2]; ●, Sa291 [class 3]). Representative data are shown.