Impact of Variations in Test Method Parameters on In Vitro Activity of Surotomycin against Clostridium difficile and Surotomycin Quality Control Limits for Broth Microdilution and Agar Dilution Susceptibility Testing

Abstract

c Test parameter variations were evaluated for their effects on surotomycin MICs. Calcium concentration was the only variable that influenced MICs; therefore, 50 g/ml (standard for lipopeptide testing) is recommended. Quality control ranges for Clostridium difficile (0.12 to 1 g/ml) and Eggerthella lenta (broth, 1 to 4 g/ml; agar, 1 to 8 g/ml) were approved by the Clinical and Laboratory Standards Institute based on these data. In recent years, the incidence and severity of Clostridium difficileassociated diarrhea (CDAD) have increased (1-4). Despite the availability of antibiotic therapies, CDAD-related morbidity and mortality rates have remained high (5-8). In a recent study, the clinical response rates were 81.1% for vancomycin and 72.7% for metronidazole, with both drugs being associated with high rates of recurrence (ϳ20 to 45%) (9-11). The more recently approved fidaxomicin demonstrated comparable clinical responses and reduced rates of recurrence (ϳ13 to 15%), as compared with vancomycin (12, 13).Surotomycin 315) is an orally administered, minimally absorbed, selective, bactericidal cyclic lipopeptide being developed for treatment of patients with CDAD (14). It previously demonstrated selective potent bactericidal activity against various C. difficile strains and other Gram-positive bacteria in vitro, with minimal impact on the Gram-negative organisms of the intestinal microbiota in vitro and in CDAD patients (15)(16)(17)(18)(19)(20). In a phase 2 trial, surotomycin was more sparing of gut microbes (primarily Bacteroides and Prevotella species) than vancomycin, with evidence for reduction in the risk of recurrence (20).These studies were designed to evaluate the effects of changes in key test parameters, including pH, inoculum, calcium concentration ([Ca 2ϩ ]), broth/agar lot, and incubation time, on the in vitro activity of surotomycin against three C. difficile strains (ATCC 700057 and the clinical isolates SID 7907612 and SID 2946830) and to generate MIC ranges for quality control (QC) organisms (C. difficile ATCC 700057, Eggerthella lenta ATCC 43055, and Staphylococcus aureus ATCC 29213). Broth microdilution and agar dilution were reported previously to be comparable and reproducible methods for testing surotomycin against C. difficile (21). Therefore, the test parameter and QC studies were performed using both methods, based on the data presented here.Variations in key test parameters were tested in duplicate against C. difficile ATCC 700057 and two clinical isolates. Brucella agar, as recommended by the Clinical and Laboratory Standards Institute (CLSI) for MIC determinations, has various concentrations of divalent cations (22). The in vitro activity of surotomycin is dependent on the [Ca 2ϩ ] of the test medium, which needs to be adjusted to a final value of 50 g/ml, based on previous observations with the similarly structured lipopeptide daptomycin (23). Inoculum size (5 ϫ 10 3 to 5 ϫ 10 5 CFU/ ml), pH (pH 6 to 8), [Ca 2ϩ ] (0 to 150 g/ml), and incubation time (24 to 72 h) were varied relati...

Effect of Surotomycin, a Novel Cyclic Lipopeptide Antibiotic, on Intestinal Colonization with Vancomycin-Resistant Enterococci and Klebsiella pneumoniae in Mice

“…Broth dilution MICs of the test antibiotics for the test organisms were determined using standard methods for testing the susceptibility of aerobic bacteria (14,15). For surotomycin, the calcium in the medium was adjusted to a final concentration of 50 mg/liter, the optimal concentration for antibacterial activity (16).…”