Comparative Bactericidal Activities of Daptomycin and Vancomycin against Glycopeptide-Intermediate Staphylococcus aureus (GISA) and Heterogeneous GISA Isolates

Abstract

Staphylococcus aureus strains from the U.S. SENTRY Antimicrobial Surveillance Program, 2002-2003, glycopeptide-intermediate S. aureus (GISA) strains, and heterogeneous GISA (hGISA) strains were used to compare bactericidal activities of daptomycin and vancomycin using MICs and minimum bactericidal concentrations. Glycopeptide-susceptible S. aureus and hGISA strains were further studied by using time-kill curves. For all isolates, the daptomycin MIC₅₀ and MIC₉₀ are four times lower and the log drops in viable counts at 6 h and 24 h are significantly greater than those for vancomycin.

Staphylococcus aureus is an important cause of serious infections in both hospitals and the community and is particularly efficient at developing resistance to antimicrobial agents (9). In the United States, the proportion of nosocomial intensive care unit S. aureus infections due to methicillin-resistant Staphylococcus aureus (MRSA) is now >50%, and infections caused by MRSA are associated with a longer hospital stay, more antibiotic administration, and higher costs (11, 3). Until recently, glycopeptides were believed to have retained activity against all S. aureus strains, and therefore, the spread of MRSA has led to increased usage of glycopeptides and hence increased selective pressure for the development of resistance (7, 14). Although the isolation of glycopeptide-intermediate S. aureus (GISA) is rare, there seems to be a more widespread prevalence of heterogeneous GISA (hGISA) (8, 17). Consequently, we are now faced with growing problems of reduced susceptibility in S. aureus (both homogeneous and heterogeneous) and the need of alternatives for treatment.

Daptomycin has recently demonstrated significantly better bactericidal activity than vancomycin against S. aureus and enterococci (15, 16) and has activity against a small number of glycopeptide-intermediate S. aureus strains and vancomycin-resistant enterococcus (1). Daptomycin is an acidic lipopeptide with a mode of action requiring calcium (2).

The aim of this study was to compare bactericidal activities of daptomycin and vancomycin against a range of glycopeptide-susceptible and intermediately resistant Staphylococcus aureus strains. Accordingly, MICs and minimum bactericidal concentrations (MBCs) were determined for four phenotypes (methicillin-susceptible S. aureus/glycopeptide-susceptible S. aureus [MSSA/GSSA], MRSA/GSSA, hGISA/MRSA, and GISA/MRSA), and time-kill curves were used to compare bactericidal activities of both antimicrobials against the more prevalent hGISA and GSSA/MRSA isolates. Daptomycin and vancomycin MICs and MBCs were determined for 11 methicillin-susceptible and glycopeptide-susceptible S. aureus isolates (from the SENTRY program, 2002-2003), 95 MRSA/GSSA isolates (SENTRY program, 2002-2003), 55 hGISA/MRSA isolates (4 isolates from the SENTRY program, 2002-2003, and 51 from BCARE), and 15 GISA/MRSA isolates (BCARE) using a standard CLSI (formerly NCCLS) broth microdilution technique (10). Bactericidal activities of vancomycin and daptomycin were also investigated for 10 GSSA and 10 hGISA strains using standard time-kill-curve techniques (6). Strains were divided according to phenotype using the previously described population analysis profile-area under the curve method (18). MICs were determined using standard CLSI broth microdilution methodology with Mueller-Hinton Broth (MHB) alone for vancomycin and MHB with an adjusted Ca²⁺ concentration (50 mg/liter) for daptomycin. Antimicrobials were used in a log 2 dilution series from 0.06 to 64 mg/liter for daptomycin and from 0.25 to 16 mg/liter for vancomycin. Inocula were prepared from direct colony suspension, and microtiter plates were inoculated with 10⁵ CFU/ml. Plates were incubated in air at 37°C for 18 h. The MIC was defined as the lowest concentration of an antimicrobial agent that prevents visible growth of a microorganism in a broth dilution susceptibility test. MBCs were determined by subculture of 100 μl of well contents on blood agar with incubation of plates in air at 37°C for 18 h. The MBC was defined as the lowest concentration of an antimicrobial agent that reduces the initial viable count by 99.9%.

Time-kill experiments were performed using MHB alone for vancomycin and Ca²⁺-supplemented MHB for daptomycin with a 10⁶-CFU/ml inoculum. Drug concentrations used were 2× and 4× the MIC of the organism tested plus an antibiotic-free control. Samples were collected at 0, 1, 2, 4, 6, and 24 h and viable counts plotted versus antibiotic concentrations. Log drops in viable counts at 6 h and 24 h were calculated, and the bactericidal effect was defined as a ≥3 log₁₀ CFU/ml decrease of the initial inoculum after 6 h and 24 h.

The in vitro activities of daptomycin and vancomycin against all phenotypes tested are shown in Tables 1 and 2. Daptomycin MICs were higher for hGISA and GISA strains than for GSSA strains. This suggests that daptomycin MICs increase with increasing vancomycin MICs, with a GISA daptomycin mean MIC of 0.91 compared to 0.48 for hGISA strains and 0.31/0.30 for GSSA strains. However, daptomycin MIC₅₀s and MIC₉₀s were four times lower than those for vancomycin for all phenotypes tested (Table 1). The daptomycin mean MIC was four times lower than the vancomycin mean MIC for all phenotypes tested. MBCs at which 50% and 90% of the strains tested were killed (MBC₅₀s and MBC₉₀s) for daptomycin were 8 and 16 times lower than those for vancomycin for all phenotypes, including hGISA and GISA strains. Daptomycin MBC/MIC ratios were significantly lower than vancomycin MBC/MIC ratios for all phenotypes (Table 2).

TABLE 1.

Activities of daptomycin and vancomycin against a collection of S. aureus strains with various vancomycin susceptibilities

Phenotype	No. of strains	Antimicrobial agent	Range of MICs	MIC50	MIC90	% Susceptible	% Resistant	Geometric mean MIC
GSSA/MSSA	11	Vancomycin	1-2	1	2	100	0	1.1
		Daptomycin	0.25-0.5	0.25	0.5	100	0	0.3
GSSA/MRSA	95	Vancomycin	0.5-2	1	2	100	0	1.2
		Daptomycin	0.12-1	0.25	0.5	100	0	0.3
hGISA	55	Vancomycin	1-4	2	4	100	0	2.4
		Daptomycin	0.12-2	0.5	1	98.2	1.8	0.5
GISA	15	Vancomycin	4-8	4	8	60	40	5.5
		Daptomycin	0.5-2	1	2	86.7	13.3	0.9

TABLE 2.

MBCs for daptomycin and vancomycin against a collection of S. aureus strains with various vancomycin susceptibilities

Phenotype	No. of strains	Antimicrobial agent	Range of MBCs	MBC:50	MBC90	Geometric mean MBC	Mean MBC/MIC ratio
GSSA/MSSA	11	Vancomycin	1-16	8	16	6.2	5.5
		Daptomycin	0.25-4	0.5	2	0.53	2.4
GSSA/MRSA	95	Vancomycin	0.50-16	4	16	4.5	3.8
		Daptomycin	0.25-4	0.5	1	0.53	1.7
hGISA	55	Vancomycin	2-32	16	32	11.7	4.9
		Daptomycin	0.25-4	1	2	0.9	1.9
GISA	15	Vancomycin	8-32	16	32	20.2	3.6
		Daptomycin	0.5-2	2	2	1.3	1.4

In time-kill curve studies, the 10 GSSA strains used had vancomycin MICs ranging from 0.5 to 1 mg/liter and daptomycin MICs ranging from 0.12 to 1 mg/liter, while 10 hGISA strains had vancomycin MICs ranging from 1 to 4 mg/liter and daptomycin MICs ranging from 0.12 to 2 mg/liter. The mean log drop (6 h) in viable counts of GSSA with daptomycin at 2× or 4× MIC was 2.9 or 2.4 times greater than that for vancomycin (Table 3). At 24 h, the mean log drop in viable counts of GSSA for daptomycin at 2× or 4× MIC was 2.6 or 2.2 times greater than that for vancomycin. With hGISA, the mean log drop (6 h) in viable counts for daptomycin at 2× or 4× MIC was 2.5 or 2.5 times greater than that for vancomycin (Table 3). At 24 h, the mean log drop in viable counts of hGISA for daptomycin at 2× or 4× MIC was 1.9 or 1.7 times greater than that for vancomycin. In all cases, the log drops in viable counts at 2× and 4× MIC were significantly greater for daptomycin than for vancomycin (P = <0.001).

TABLE 3.

Log drop in viable count at 6 h or 24 h for 10 GSSA and 10 hGISA strains using vancomycin or daptomycin at 2× or 4× MIC^a

Phenotype	Antimicrobial concnb	Range of log drop in viable count (0-6 h)	Mean log drop in viable count (0-6 h)	Range of log drop in viable count (0-24 h)	Mean log drop in viable count (0-24 h)
GSSA	V × 2 MIC	0.42-1.8	1.1	0-2.8	1.5
	V × 4 MIC	1.1-2.4	1.5	1.5-3	2.3
	D × 2 MIC	2.1-6	3.1	1.8-6	3.8
	D × 4 MIC	2.4-5.9	3.7	2.3-6	4.9
hGISA	V × 2 MIC	0.28-3.2	1.3	0.07-3.8	2.2
	V × 4 MIC	0.43-3.5	1.4	1.2-4.5	3.4
	D × 2 MIC	1.9-6	3.2	3.3-6	4.3
	D × 4 MIC	2.4-6	3.5	4.3-6	5.7

^aLog drop assumes limit of 6× log₁₀.

^bV × 2 or V × 4 MIC, vancomycin at 2× or 4× MIC, respectively; D × 2 or D × 4 MIC, daptomycin at 2× or 4× MIC, respectively.

At 6 h, daptomycin was bactericidal (≥3-log drop) for 50% of GSSA strains at 2× MIC and for 70% of GSSA strains at 4× MIC compared with no GSSA strains at both 2× and 4× MIC for vancomycin. With hGISA, daptomycin was bactericidal at 6 h for 30% of strains at 2× MIC and 60% of strains at 4× MIC compared with 10% of strains at both 2× and 4× MICs for vancomycin. At 24 h, daptomycin was bactericidal for 70% of GSSA strains at 2× MIC and 90% of strains at 4× MIC compared with 10% of GSSA strain at 4× MIC for vancomycin. For hGISA, daptomycin was bactericidal at 24 h for 100% of strains at both 2× and 4× MIC compared with 20% of strains at 2× MIC and 80% of strains at 4× MIC for vancomycin.

Bactericidal activity is probably essential for effective treatment of high-bacterial-density infections, such as bacterial endocarditis and serious infections in immunocompromised patients (5). In addition, the emergence of reduced susceptibility to glycopeptides and in particular the more highly prevalent hGISA has increased pressure for effective treatment options. In this study, MIC/MBC data confirm those of previous studies, which show higher daptomycin MICs for some strains with reduced susceptibility to vancomycin (4, 13). This suggests that the development of heterogeneous vancomycin resistance and more specifically the thickening of the bacterial cell wall may act as a barrier to the large daptomycin molecule (4). However, bactericidal activity, as determined by MBC₅₀s, MBC₉₀s, and MBC/MIC ratios, shows that daptomycin is considerably more bactericidal than vancomycin against glycopeptide-susceptible, hGISA, and GISA strains. Time-kill curves also clearly show that daptomycin is significantly more bactericidal for each strain, especially at 6 h, than vancomycin for both GSSA and hGISA. Daptomycin was also bactericidal against one daptomycin-resistant hGISA strain (MIC of 2 mg/liter). These data confirm that daptomycin shows bactericidal activity against hGISA and suggest that the bactericidal activity of daptomycin is affected little by the decreased vancomycin susceptibility seen with hGISA (12).

In summary, the data presented here show that despite the slightly raised MICs seen for strains with reduced susceptibility to vancomycin, daptomycin has greater bactericidal activity than vancomycin for hGISA and GISA and can be considered a valid alternative to vancomycin in the treatment of infections caused by MRSA, hGISA, or GISA.