Efficacy of High Doses of Daptomycin versus Alternative Therapies against Experimental Foreign-Body Infection by Methicillin-Resistant Staphylococcus aureus

Since the currently approved dose of daptomycin (6 mg/kg of body weight/day) has been associated with clinical failures and resistance development, higher doses for some difficult-to-treat infections are being proposed. We studied the efficacy of daptomycin at high doses (equivalent to 10 mg/kg/day in humans) and compared it to that of reference and alternative treatments in a model of foreign-body infection with methicillin (meticillin)-resistant Staphylococcus aureus. In vitro studies were conducted with bacteria in the log and stationary phases. For the in vivo model, therapy with daptomycin at 100 mg/kg/day, vancomycin at 50 mg/kg/12 h, rifampin (rifampicin) at 25 mg/kg/12 h, or linezolid at 35 mg/kg/12 h was administered for 7 days. Antibiotic efficacy was evaluated using either bacteria from tissue cage fluids or those attached to coverslips. We screened for the emergence of linezolid- and rifampin-resistant strains and analyzed the surviving population from the daptomycin-treated group. Only daptomycin was bactericidal in both the log- and stationary-phase studies. Daptomycin (decrease in the log number of CFU per milliliter of tissue cage fluid, 2.57) and rifampin (decrease, 2.6 log CFU/ml) were better (P < 0.05) than vancomycin (decrease, 1.1 log CFU/ml) and linezolid (decrease, 0.9 log CFU/ml) in the animal model. Rifampin-resistant strains appeared in 60% of cases, whereas no linezolid resistance emerged. No daptomycin-resistant subpopulations were detected at frequencies of 10⁻⁷ or higher. In conclusion, daptomycin at high doses proved to be as effective as rifampin, and the two were the most active therapies for this experimental foreign-body infection. These high doses ensured a profile of safety from the development of resistance.Daptomycin is a lipopeptide drug with bactericidal activity toward methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) in a concentration-dependent manner (27, 32). It is currently approved for use at 4 mg/kg of body weight/day for skin and soft-tissue infections (1) and at 6 mg/kg/day for bacteremia and right-side endocarditis (12, 14).In recent years, reports of clinical failures and the emergence of resistant strains following daptomycin treatment have raised great concern (6, 18, 31). As a result, higher doses of daptomycin are being proposed as an alternative for some difficult-to-treat infections such as complicated bacteremia and endocarditis. Recently, doses of 10 mg/kg/day were studied using an in vitro model of staphylococcal endocarditis, with the results being promising in terms of efficacy and resistance prevention (25, 26). To date, clinical experience with the activity of the drug at doses higher than 6 mg/kg/day is limited (15, 28), whereas good safety and tolerance profiles for daptomycin at up to 12 mg/kg/day in volunteers have been reported (4).Foreign-body infections are difficult to treat because of the presence of bacterial biofilm and tolerance to antibiotics (10, 39). MRSA is commonly involved in such infections, and daptomycin may be a promising drug (39). However, clinical experience in this area is again scarce, and the recommended doses are not clearly established, there being reports of clinical failures with doses of 4 to 6 mg/kg/day (13, 24).The rat model of tissue cage infection is a well-standardized model that reasonably mimics human device infections (17, 20, 35). In this model, the efficacy of daptomycin has been partially studied (29, 34).Taken together, the available experimental and clinical data seem to indicate that high doses of daptomycin are required in the setting of foreign-body infection.In the present study, we aimed to test the efficacy of daptomycin at doses equivalent to 10 mg/kg/day in humans in a model of foreign-body infection with MRSA, comparing it with the efficacy of the current reference or main alternative treatments such as vancomycin, rifampin (rifampicin), and linezolid. We also sought to analyze the protection offered against the emergence of resistant strains.     

Fosfomycin-Daptomycin and Other Fosfomycin Combinations as Alternative Therapies in Experimental Foreign-Body Infection by Methicillin-Resistant Staphylococcus aureus

The efficacy of daptomycin, imipenem, or rifampin with fosfomycin was evaluated and compared with that of daptomycin-rifampin in a tissue cage model infection caused by methicillin-resistant Staphylococcus aureus (MRSA). Strain HUSA 304 was used. The study yielded the following results for MICs (in μg/ml): fosfomycin, 4; daptomycin, 1; imipenem, 0.25; and rifampin, 0.03. The study yielded the following results for minimum bactericidal concentration (MBC) (in μg/ml): fosfomycin, 8; daptomycin, 4; imipenem, 32; and rifampin, 0.5. Daptomycin-rifampin was confirmed as the most effective therapy against MRSA foreign-body infections. Fosfomycin combinations with high doses of daptomycin and rifampin were efficacious alternative therapies in this setting. Fosfomycin-imipenem was relatively ineffective and did not protect against resistance.     

High Activity of Fosfomycin and Rifampin against Methicillin-Resistant Staphylococcus aureus Biofilm In Vitro and in an Experimental Foreign-Body Infection Model

Increasing antimicrobial resistance reduces treatment options for implant-associated infections caused by methicillin-resistant Staphylococcus aureus (MRSA). We evaluated the activity of fosfomycin alone and in combination with vancomycin, daptomycin, rifampin, and tigecycline against MRSA (ATCC 43300) in a foreign-body (implantable cage) infection model. The MICs of the individual agents were as follows: fosfomycin, 1 μg/ml; daptomycin, 0.125 μg/ml; vancomycin, 1 μg/ml; rifampin, 0.04 μg/ml; and tigecycline, 0.125 μg/ml. Microcalorimetry showed synergistic activity of fosfomycin and rifampin at subinhibitory concentrations against planktonic and biofilm MRSA. In time-kill curves, fosfomycin exhibited time-dependent activity against MRSA with a reduction of 2.5 log₁₀ CFU/ml at 128 × the MIC. In the animal model, planktonic bacteria in cage fluid were reduced by <1 log₁₀ CFU/ml with fosfomycin and tigecycline, 1.7 log₁₀ with daptomycin, 2.2 log₁₀ with fosfomycin-tigecycline and fosfomycin-vancomycin, 3.8 log₁₀ with fosfomycin-daptomycin, and >6.0 log₁₀ with daptomycin-rifampin and fosfomycin-rifampin. Daptomycin-rifampin cured 67% of cage-associated infections and fosfomycin-rifampin cured 83%, whereas all single drugs (fosfomycin, daptomycin, and tigecycline) and rifampin-free fosfomycin combinations showed no cure of MRSA cage-associated infections. No emergence of fosfomycin resistance was observed in animals; however, a 4-fold increase in fosfomycin MIC (from 2 to 16 μg/ml) occurred in the fosfomycin-vancomycin group. In summary, the highest eradication of MRSA cage-associated infections was achieved with fosfomycin in combination with rifampin (83%). Fosfomycin may be used in combination with rifampin against MRSA implant-associated infections, but it cannot replace rifampin as an antibiofilm agent.     

Comparison of Tigecycline and Vancomycin for Treatment of Experimental Foreign-Body Infection Due to Methicillin-Resistant Staphylococcus aureus

Twice-daily 7-day regimens of tigecycline (7 mg/kg) and vancomycin (50 mg/kg) were compared in a rat tissue cage model of chronic foreign-body infection due to methicillin (meticillin)-resistant Staphylococcus aureus strain MRGR3. Subcutaneously administered tigecycline reached levels in tissue cage fluid that were nearly equivalent or slightly superior to the antibiotic MIC (0.5 μg/ml) for strain MRGR3. After 7 days, equivalent, significant reductions in bacterial counts were recorded for tigecycline-treated and vancomycin-treated rats, compared with those for untreated animals.Antimicrobial therapy for foreign-body infections due to Staphylococcus aureus is challenging (38), in particular for multidrug-resistant hospital-associated and community-acquired isolates of methicillin (meticillin)-resistant S. aureus (MRSA) (3, 12, 15, 16). Tigecycline is a novel injectable glycylcycline broad-spectrum antibiotic that demonstrates excellent in vitro and in vivo activity against MRSA and other multiresistant organisms (9, 11, 22, 28, 32) and can overcome both major tetracycline resistance mechanisms, namely ribosomal protection (10, 23) and efflux (4, 27). Tigecycline has shown good activity in various animal models of serious MRSA infections (21, 39, 40), as well as against biofilm-embedded bacteria (14, 26).We previously used a rat tissue cage model of S. aureus chronic foreign-body infections for evaluating a number of antimicrobial agents, namely vancomycin (17), teicoplanin (31), imipenem (30), ceftobiprole (37), daptomycin (29, 35), and several fluoroquinolones (2, 17, 36). This study reports the activity of tigecycline compared to that of the reference anti-MRSA agent vancomycin in a tissue cage model of MRSA chronic foreign-body infection.(This study was presented in part at the 18th European Congress of Clinical Microbiology and Infectious Diseases, Barcelona, Spain, April 2008.)MRSA strain MRGR3, whose properties were previously described (2, 5, 17, 29-31, 36, 37), was used for in vitro and in vivo studies. Strain MRGR3 is resistant to methicillin, gentamicin, erythromycin, tetracycline, and chloramphenicol (17).MICs of freshly prepared (1, 13, 25) tigecycline (Wyeth Research, Collegeville, PA) or vancomycin (Vancocin; Teva Pharma AG, Switzerland) for MRSA strain MRGR3 or quality control S. aureus ATCC 29213 were determined by broth macrodilution in cation-adjusted Mueller-Hinton broth (CAMHB), according to Clinical and Laboratory Standards Institute guidelines (7).The animal protocol used for evaluating the in vivo activities of tigecycline and vancomycin was previously described in detail (17, 37) and approved by the Ethics Committee of the Faculty of Medicine, University of Geneva, and the Veterinary Office of the State of Geneva. Three weeks after subcutaneous implantation of four tissue cages per animal in anesthetized Wistar rats (37), tissue cage fluids were checked for sterility (17).Pilot pharmacokinetic studies were performed using groups of noninfected rats to find an adequate dosing regimen of tigecycline for therapy of tissue cage infections as described previously (37). Tigecycline levels in cage fluids (and blood) were estimated by a microbiological assay (21), with a detection limit of 0.25 μg/ml. To account for protein binding, all plasma or tissue cage fluid samples were diluted with 1 volume of phosphate-buffered saline and assayed in duplicate, with reference to duplicate standard concentrations (0.25 to 8 μg/ml) of tigecycline, in phosphate-buffered saline supplemented with 50% plasma or pooled tissue cage fluids, respectively.Each tissue cage was chronically infected by inoculating 5 × 10⁵ CFU of log-phase MRGR3 (37). Two weeks later, all rats whose cage fluids contained ≥10⁵ CFU/ml received twice-daily doses (by the subcutaneous route for 7 days) of tigecycline (7 mg/kg), vancomycin (50 mg/kg), or no antibiotic (control group). Differences in CFU counts of cage fluid quantitative cultures, performed at day 1 (before treatment) and day 8 (12 h after the last injection of either tigecycline or vancomycin), were expressed as the change in number of log₁₀ CFU/ml (37) and evaluated by one-way analysis of variance and post-analysis of variance pairwise comparisons between individual groups via the Tukey HSD test (http://faculty.vassar.edu/lowry/VassarStats.html), using P values of <0.05 with two-tailed significance levels.Tigecycline resistance was screened by plating 10-fold-diluted cage fluids (100 μl) onto MH agar supplemented with 2 μg/ml tigecycline. No single colony grew on tigecycline-supplemented plates inoculated with 10⁸ CFU of in vitro-grown cultures of strain MRGR3.The MIC of tigecycline in CAMHB for MRSA strain MRGR3 was 0.5 μg/ml, namely at the upper limit of susceptibility breakpoints (7), and was unaffected by supplementation of CAMHB with 50% tissue cage fluid (data not shown). Since tigecycline did not produce a 3-log₁₀ reduction in the number of MRGR3 CFU/ml, it was not considered bactericidal. Nevertheless, supra-MIC levels (1, 2, and 4 μg/ml) of tigecycline produced a 2- to 3-log₁₀ decrease in the number of MRGR3 CFU/ml at 24 h. The vancomycin MIC and minimal bactericidal concentration for strain MRGR3 were 1 and 2 μg/ml, respectively (17).Average tigecycline levels, scored for tissue cage fluids (n = 6) from 0 to 12 h after subcutaneous administration, remained quite constant over time, showing ≤3-fold variations between results at different time points and moderate animal-to-animal differences (Fig. ​(Fig.1).1). A 7-mg/kg twice-daily regimen yielded cage fluid levels of 0.39 to 0.70 μg/ml tigecycline at day 4 and 0.33 to 1.01 μg/ml at day 7, such results thus being nearly equivalent or slightly superior to the antibiotic MIC for MRGR3. Tigecycline plasma levels at 2 h on day 4 were 1.87 ± 0.66 μg/ml, in agreement with other reports (8, 21). A 14-mg/kg twice-daily regimen led to plasma and tissue cage fluid tigecycline levels ca. twofold higher than the 7-mg/kg regimen (Fig. ​(Fig.1).1). Average peak and trough cage fluid levels of vancomycin were previously determined (17) as 12 and 2 μg/ml at 4 and 12 h, respectively.Open in a separate windowFIG. 1.Pharmacokinetic levels of tigecycline in tissue cage fluids of rats on day 4 (open symbols) or day 7 (closed symbols) of therapy every 12 h with 7 mg/kg (○) or 14 mg/kg (▵) of tigecycline. Each value is the mean result of six determinations.At day 1, mean bacterial counts for MRGR3-infected cages were not significantly different (P = 0.65) in controls (6.85 ± 0.19 log₁₀ CFU/ml; n = 28), tigecycline-treated rats (6.92 ± 0.13 log₁₀ CFU/ml; n = 29), or vancomycin-treated rats (6.70 ± 0.18 log₁₀ CFU/ml; n = 27). At day 8, significant (P < 0.01 versus controls) reductions were recorded in bacterial counts in cage fluids of both tigecycline-treated (−0.62 ± 0.17 CFU/ml; n = 29) and vancomycin-treated (−0.76 ± 0.18 log₁₀ CFU/ml; n = 27) rats, whereas the bacterial counts for controls slightly increased (+0.18 ± 0.19 log₁₀ CFU/ml; n = 28) (Fig. ​(Fig.2).2). The reductions in CFU counts for vancomycin-treated and tigecycline-treated rats were not significantly different. Finally, no MRGR3 isolate showing increased tigecycline MIC was observed in any posttherapy cage fluid sample (n = 29). The lack of emergence of MRGR3 derivates with diminished susceptibility to tigecycline is consistent with the difficulty in selecting laboratory-derived, tigecycline-resistant mutants of S. aureus (18), and it contrasts with the emergence of resistant subpopulations during low-dose daptomycin therapy of S. aureus-infected tissue cages (35).Open in a separate windowFIG. 2.Decrease in viable counts of MRSA MRGR3 in tissue cage fluids of rats treated for 7 days with tigecycline or vancomycin.Several studies performed with the rat tissue cage model demonstrated the low initial in vivo response of foreign-body-associated chronic MRSA infections (2, 5, 6, 17, 20, 29-31, 35-37). A much greater reduction of viable MRSA counts in cage fluids requires longer periods of antibiotic therapy (5), as found in clinical situations with foreign-body infections (38). Major pharmacokinetic properties of tigecycline, observed in human and animal studies, are very low plasma levels, long half-lives, and high volumes of distribution indicating extensive tigecycline distribution into the tissues (8, 11, 19, 28, 32, 40). In line with previous observations that showed a requirement for active, preferentially bactericidal, antibiotic levels for obtaining significant reductions of CFU counts in MRSA-infected cage fluids (29, 37), we selected for therapy a twice-daily 7-mg/kg regimen yielding cage fluid tigecycline levels above the MIC for strain MRGR3 for >50% of the dosing interval (32, 33), while minimizing the occurrence of side effects previously observed with higher-dose regimens (39). Our regimen is similar to those required for activity in other animal models of hard-to-treat S. aureus infections, such as endocarditis or osteomyelitis (21, 39), although its relevance to human therapy is not fully defined (32). In addition, the incomplete in vitro killing activity of tigecycline, namely a <3-log₁₀ reduction in number of MRGR3 CFU at 24 h, prevents a pharmacodynamic analysis of tigecycline in vivo activity more detailed than those of previously evaluated bactericidal antibiotics in MRSA-infected cages (29, 37). We can also speculate that other properties of tigecycline, namely its in vivo activity against intracellular, slowly growing, or biofilm-forming bacteria, might significantly contribute to tigecycline activity in MRSA-infected cages (34). Indeed, high intracellular levels of tigecycline were shown to accumulate in human polymorphonuclear neutrophils and prevent growth of phagocytized bacteria (24). Further studies are needed to elucidate the mechanisms of tigecycline activity against hard-to-treat MRSA infections.     

Comparative Efficacies of Telavancin and Vancomycin in Preventing Device-Associated Colonization and Infection by Staphylococcus aureus in Rabbits

Telavancin is an investigational lipoglycopeptide antibiotic that is active against gram-positive pathogens. In an in vivo rabbit model, subtherapeutic (15-mg/kg) and therapeutic (30- or 45-mg/kg) doses of telavancin were demonstrated to be noninferior and superior to vancomycin (20 mg/kg), respectively, for preventing subcutaneous implant colonization and infection by Staphylococcus aureus.Estimated annual infection rates in the United States associated with the most commonly used medical devices range from 3% to 8% for central venous catheters, 10% to 30% for bladder catheters, and 5% to 10% for fracture fixation devices (2). Since more than 37 million of these devices are inserted annually, device-associated infections affect millions of patients and, as such, are a major medical and economic issue (2).Telavancin is an investigational lipoglycopeptide, with activity against clinically relevant gram-positive pathogens, including Staphylococcus aureus (8, 9, 15), which is one of the most important bacterial species implicated in the pathogenesis of device-related infections (2). In clinical trials, telavancin has been shown to be efficacious for the treatment of complicated skin and skin structure infections (17-19) as well as hospital-acquired pneumonia (14). The present study provides preclinical evidence that telavancin may also be more efficacious for the prevention of device colonization and infection by gram-positive pathogens than vancomycin. We used vancomycin rather than a β-lactam antibiotic as a control prophylactic agent for the following three reasons. (i) Vancomycin is currently the most commonly used agent for perioperative systemic prophylaxis when inserting surgical implants, regardless of the patient''s status of methicillin-resistant Staphylococcus aureus colonization. (ii) Vancomycin has been reported to be superior to some β-lactam antibiotics in preventing certain postoperative infections. For instance, in a randomized, double-blinded trial by Maki and colleagues, the preoperative prophylactic use of vancomycin in patients embarking on cardiac and vascular operations was more protective against infection than the use of cefazolin or cefamandole. As a result, the authors suggested the use of vancomycin as an antibiotic prophylaxis in prosthetic valve replacement and prosthetic vascular graft implantation to reduce the risk of implant infection (12). (iii) Although both vancomycin and β-lactam antibiotics share with telavancin a rather similar mechanism of action (inhibition of cell wall synthesis), it was important to determine if the two structurally related glycopeptide compounds, vancomycin and telavancin, indeed differ in their efficacies.This study was conducted with prior approval from the appropriate Institutional Animal Care and Use Committee (IACUC). Modifications to a previously described rabbit model of subcutaneous implant colonization and infection by S. aureus were used in these studies (3, 4, 6). In a similar rabbit model that compared vancomycin to dalbavancin, a compound closely related to telavancin, we demonstrated a statistically insignificant trend for lower rates of device colonization in the dalbavancin group than in the vancomycin group. Telavancin was obtained from Theravance, Inc. (South San Francisco, CA), vancomycin from Hospira (Lake Forest, IL), and water with 5% dextrose (D5W) from IVX Animal Health (St. Joseph, MO). In vitro bacterial susceptibility to telavancin and vancomycin was tested, in triplicate, by standard macrodilution (1). We used specific-pathogen-free, 4- to 7-month-old female New Zealand White rabbits, with a body mass of 3 to 4 kg each (Myrtle''s Rabbitry, Thomson Station, TN). Anesthesia was induced by intramuscular injection of xylazine at 6 mg/kg and acepromazine at 2 mg/kg and maintained via inhalation of 0.5% to 2% isoflurane for the duration of the surgery.Anesthetized animals were randomly assigned to one of five treatment groups (nine rabbits each) and injected, intravenously, over a period of ∼2 min, with a single dose of sterile D5W (control), vancomycin (20 mg/kg), or one of three doses of telavancin (15, 30, or 45 mg/kg). Based on previously reported efficacy and exposure data for rabbits (11), this telavancin dose range of 15 to 45 mg/kg for rabbits corresponds to 50% to 150% of the human area under the concentration-time curve-equivalent dose (HED) of 10 mg/kg (10), which is also the recommended human clinical dose (18). The vancomycin dose used in this study was similar to those used in prior rabbit studies, including this particular model (3). Following a surgical procedure described previously (3), six 2-cm-long segments of seven French triple-lumen polyurethane vascular catheters were implanted subcutaneously in the back of each animal, for a total of 54 devices per nine rabbits in each group. The implanted devices were inoculated on the surface directly with 10⁵ CFU (50-μl total inoculum in Trypticase soy broth) using strain P1 of S. aureus, a methicillin-sensitive S. aureus (MSSA) clinical isolate associated with device-related infections that has been used by us and others for studies of medical device colonization (3, 6). The mean MIC and minimal bactericidal concentration of the MSSA P1 strain used in this study were 0.25 μg/ml and 1 μg/ml for telavancin, respectively, and 1 μg/ml and 8 μg/ml for vancomycin, respectively. Surgical wounds were closed, and the animals were observed closely in the operating room until they achieved sternal recumbency. The analgesic/anti-inflammatory agent ketoprofen (3 mg/kg) was given intramuscularly to each rabbit immediately following surgery and as needed thereafter. The animals were monitored daily for signs of pain, distress, erythema, local infection, and sepsis.One week postsurgery, all rabbits were anesthetized and humanely sacrificed. Implanted catheter segments were recovered in a sterile fashion and cultured based on a previously described sonication technique (13, 16). Swab cultures were collected from the soft tissues adjacent to the implantation site or wound drainage, while blood samples were collected by cardiac puncture. The surgical site swabs as well as the blood samples were cultured using standard techniques (4, 5).Device colonization was defined as growth of the inoculated S. aureus strain from the sonication culture of the explanted device. Device-related infection was defined as any growth of the inoculated strain from both the sonication culture of the explanted device and the swab culture of any soft tissue collection or wound site discharge. Explanted devices were sonicated in 2 ml of normal saline, and 200 μl of the sonicate and subsequent dilutions were cultured. As a result, the detectability limit was 10 CFU.The sample size was determined based on our previous experience with this animal model, which showed a 19% reduction in the rate of infection when using dalbavancin rather than vancomycin (3). In this study, we sought a similar magnitude of reduction (19%) in the frequency of infection when using telavancin compared to vancomycin with a power of 80% and a type I error of 5%. Frequencies of device colonization and local device-related infections in the five treatment groups were compared using a two-tailed Fisher''s exact test at an alpha level of 0.05 (Stata statistical software, version 8.2; StataCorp, College Station, TX).The mean MIC and minimal bactericidal concentration of the MSSA P1 strain used in this study were 0.25 μg/ml and 1 μg/ml for telavancin, respectively, and 1 μg/ml and 8 μg/ml for vancomycin, respectively. Blood cultures derived from all groups were sterile, indicating the absence of device-related bacteremia. The frequencies of device colonization and device-associated infection by the inoculated S. aureus strain were significantly higher in the control group than in each of the four treatment groups (Table ​(Table1).1). Preoperative systemic administration of telavancin reduced the rates of S. aureus device colonization and device-associated infection in a dose-related fashion. A subtherapeutic dose of telavancin (15 mg/kg), representing a HED of 4.8 mg/kg, was noninferior to vancomycin at 20 mg/kg in preventing device colonization and device-related infections. Single therapeutic doses of telavancin (30 mg/kg or 45 mg/kg), representing HEDs of 9.6 and 14.4 mg/kg, respectively, were superior to vancomycin (20 mg/kg) at preventing device colonization and device-related infections (Table ​(Table11).

TABLE 1.

Rates of device colonization and device-related infection by S. aureus

Vancomycin-Rifampin Combination Therapy Has Enhanced Efficacy against an Experimental Staphylococcus aureus Prosthetic Joint Infection

Treatment of prosthetic joint infections often involves a two-stage exchange, with implant removal and antibiotic spacer placement followed by systemic antibiotic therapy and delayed reimplantation. However, if antibiotic therapy can be improved, one-stage exchange or implant retention may be more feasible, thereby decreasing morbidity and preserving function. In this study, a mouse model of prosthetic joint infection was used in which Staphylococcus aureus was inoculated into a knee joint containing a surgically placed metallic implant extending from the femur. This model was used to evaluate whether combination therapy of vancomycin plus rifampin has increased efficacy compared with vancomycin alone against these infections. On postoperative day 7, vancomycin with or without rifampin was administered for 6 weeks with implant retention. In vivo bioluminescence imaging, ex vivo CFU enumeration, X-ray imaging, and histologic analysis were carried out. We found that there was a marked therapeutic benefit when vancomycin was combined with rifampin compared with vancomycin alone. Taken together, our results suggest that the mouse model used could serve as a valuable in vivo preclinical model system to evaluate and compare efficacies of antibiotics and combinatory therapy for prosthetic joint infections before more extensive studies are carried out in human subjects.     

Bactericidal Activity of the Combination of Levofloxacin with Rifampin in Experimental Prosthetic Knee Infection in Rabbits Due to Methicillin-Susceptible Staphylococcus aureus

The combination of levofloxacin and rifampin has been recommended for the treatment of staphylococcal prosthetic infection. In a rabbit model of prosthetic knee infection due to a susceptible clinical strain of Staphylococcus aureus, the combination of levofloxacin and rifampin was bactericidal, significantly reduced bacterial titers in bone compared with levels for rifampin and controls (P < 0.05), sterilized 6 of 12 animals, and prevented the selection of resistant mutants that was observed with rifampin alone, validating clinical recommendations.Because of its pharmacodynamic and pharmacokinetic properties, rifampin is considered a cornerstone of antibiotic treatment of staphylococcal prosthesis joint infections (14, 19). Indeed, rifampin is highly active in vitro against staphylococci, has good diffusion in bone, is active against small-colony variants sequestered into phagocytic cells (16), and retains bactericidal activity against nongrowing staphylococci, such as those that adhere to foreign bodies (11, 19). Rifampin has been shown to be effective in experimental studies (8, 14). However, this drug should not be given alone, due to the frequent development of resistant mutants (19). Thus, the combination of rifampin with fluoroquinolones that share pharmacodynamic and pharmacokinetic properties with rifampin has been shown to be the most effective treatment for such infections (4, 8, 14, 19). Levofloxacin, the l-isomer of the racemate ofloxacin, has a high level of in vitro activity against Staphylococcus aureus (7) and demonstrated good activity in foreign-body experimental models (13, 17) and good penetration in bone samples from patients undergoing arthroplasty (12). Levofloxacin has been recommended for treatment of staphylococcal prosthetic joint infections in adults (20). However, its specific activity in prosthetic joint infections has not been clearly established. We therefore evaluated the efficacy of levofloxacin, alone or in combination with rifampin, for treatment of rabbit experimental prosthetic knee infections due to S. aureus.(This work was presented in part at the 46th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, 2006.)S. aureus 17848, isolated from a patient with an infected knee prosthesis, was used for all experiments. The MICs of levofloxacin and rifampin were determined by using the Etest method (AB Biodisk, Solna, Sweden) as recommended by the manufacturer. The mutant prevention concentration (MPC), recorded as the lowest antibiotic concentration completely inhibiting bacterial growth after incubation at 37°C for 72 h, was determined for rifampin and levofloxacin, as described previously (2). Time-kill curve studies were used to determine the bactericidal activities of levofloxacin and rifampin alone and in combination. Exponential-phase cultures were diluted in 10 ml of fresh Mueller-Hinton broth to yield a final inoculum of 10⁶ CFU/ml. The concentrations of antibiotic used were equivalent to 1× MIC and 4× MIC for levofloxacin and 2× MIC for rifampin. After 0, 3, 6, and 24 h of incubation in a shaking water bath at 37°C, serial dilutions of 0.1-ml samples were subcultured onto trypticase soy agar plates and incubated at 37°C for 24 h before CFU were counted. Bactericidal effect was defined by a ≥3-log₁₀ decrease of the initial inoculum. Synergy was defined by a decrease of at least 2 log₁₀ CFU/ml between the combination and its most active constituent after 24 h of incubation.New Zealand White rabbits, each weighing between 2.5 and 3 kg, were experimentally infected. Animal experiments were performed in accordance with prevailing regulations in the European Commission (9). This model has been described in detail elsewhere (1, 5). Briefly, a silicone-elastomer implant, commonly used in arthroplasty of the first metatarsophalangeal joint (Silastic HP great toe implant; Swanson Design, Dow-Corning [provided by Ortho Technique, Créteil, France]) was implanted as a tibial prosthetic component under general anesthesia. Immediately after surgery, animals were inoculated with of 10⁷ CFU of S. aureus in a final volume of 0.5 ml in phosphate-buffered saline, injected into the knee close to the prosthesis.Seven days after inoculation (day 7), rabbits were randomly assigned to an untreated control group (n = 10) or a group receiving treatment with levofloxacin alone (n = 12), rifampin alone (n = 11), or levofloxacin combined with rifampin (n = 12). Each regimen was administered for 7 days. Antibiotic dosing regimens were chosen in order to reproduce plasma concentrations in humans. For rifampin, a dosing regimen of 10 mg/kg of body weight twice a day, given intramuscularly, was used, as it produced peak levels of 9.3 ± 0.5 μg/ml in plasma 2 h after injection, which were close to the levels obtained for humans after a 10-mg/kg dose (18). The levofloxacin dosing regimen was determined in pilot studies in order to obtain peak concentrations and areas under the concentration-time curve over 24 h (AUC_0-24 levels) comparable to those achieved for humans with a dose of 750 mg per day, given intravenously (i.v.), i.e., peak levels of 11 to 12 μg/ml and AUC_0-24 levels of 90 μg·h/ml (3, 7). The dosing regimen of levofloxacin tested which best reproduced human levels was 25 mg/kg twice a day, given i.v., which produced peak levels ranging from 12.8 to 14.4 μg/ml and AUC_0-24 levels ranging from 58 to 78 μg·h/ml 15 min after injection.All the animals were killed by i.v. injection of pentobarbital 3 days after the end of therapy (day 17). The upper third (length, 3 cm) of the tibia, including compact bone and marrow, was isolated, split with a bone crusher, weighed, cut into little pieces, frozen in liquid nitrogen, and then crushed in an autopulverizer (Spex 6700 freezer; Mill Industries, Inc., NJ). The pulverized bone was suspended into 10 ml of sterile saline. Serial dilutions were made, and 0.1 ml of the dilution was plated on trypticase soy agar and horse blood agar plates. After 48 h of incubation at 37°C, the number of viable organisms was determined. The lowest detectable bacterial counts according to bone weight were 1.53 to 1.83 log₁₀ CFU/g bone. Mutant-resistant S. aureus isolates were detected in bone samples from six untreated rabbits randomly chosen and in all the treated rabbits. Portions (0.1 ml) of undiluted bone homogenate were plated onto Mueller-Hinton agar containing levofloxacin at a concentration of twofold the MIC or rifampin at a concentration of eightfold the MIC. After 72 h of incubation at 37°C, colonies were detected and MICs of levofloxacin and rifampin were measured.Peak and trough levofloxacin plasma levels were determined for six infected rabbits 10 min (peak) and 12 h (trough) after i.v. injection on the last day of therapy for animals treated with levofloxacin alone. The levofloxacin and rifampin concentrations in bone samples from all treated animals were determined using 1-ml portions of the pulverized bone. Levofloxacin and rifampin concentrations were measured by high-performance liquid chromatography.The MICs of levofloxacin and rifampin were 0.125 and 0.016 μg/ml, respectively, and the MPCs were 1 and 256 μg/ml, respectively. The results for the time-kill studies are presented in Fig. ​Fig.1.1. Rifampin alone at a concentration of 2× MIC was associated with a bacterial regrowth at 24 h after an initial bacterial killing. Levofloxacin alone exhibited a concentration-dependent killing, with a bacteriostatic effect followed by a regrowth after 24 h of incubation at 1× MIC, compared with a rapid bactericidal activity at a concentration of 4× MIC. The combination of rifampin and levofloxacin prevented the regrowth observed with rifampin alone at both concentrations of levofloxacin tested. The combination of rifampin with levofloxacin (1× MIC) was synergistic. In contrast, with a higher concentration of levofloxacin (4× MIC), the bactericidal activity of the combination tended to be reduced compared with that of levofloxacin alone after 3 and 6 h of exposure, without achieving antagonism.Open in a separate windowFIG. 1.Time-kill curve study of Staphylococcus aureus 17848 incubated for 24 h in Mueller-Hinton broth with no antibiotic (Control), levofloxacin at a concentration equal to the MIC (Lev, 1× MIC), levofloxacin at a concentration equal to 4× MIC (Lev, 4× MIC), rifampin at a concentration equal to 2× MIC (Rif, 2× MIC), a combination of levofloxacin at a concentration equal to the MIC and rifampin at a concentration equal to 2× MIC (Lev, 1× MIC + Rif), and combination of levofloxacin at a concentration equal to 4× MIC and rifampin at a concentration equal to 2× MIC (Lev, 4× MIC + Rif).The levofloxacin concentrations in plasma from infected rabbits obtained 15 min and 12 h after the end of the infusion were 17.3 ± 3.3 μg/ml and 0.29 ± 0.13 μg/ml, respectively. In bone samples, 72 h after the end of therapy, the concentrations of levofloxacin were 0.18 ± 0.06 μg/g (range, 0.09 to 0.29 μg/g), whereas the concentrations of rifampin were undetectable in seven animals and ranged from 0.15 to 7.95 μg/g in the remaining 16 rabbits.The results obtained with the different therapeutic regimens tested are shown in Table ​Table1.1. Rifampin alone significantly reduced bacterial titers in bone compared with levels for untreated controls (P < 0.05) and sterilized 5 of 11 animals. As expected, four of the six animals treated with rifampin alone that were not sterilized retained mutant-resistant strains. This result can easily be explained by the high rifampin MPC (256 μg/ml) that could not be achieved by local concentrations of rifampin in bone (<8 μg/g in all cases). Levofloxacin alone was bactericidal, significantly reduced bacterial titers in bone compared with levels for untreated controls (P < 0.05), and sterilized 5 of 12 animals (42%). No mutants resistant to levofloxacin were detected in bone samples from animals that were not sterilized at the time of sacrifice.

TABLE 1.

Effects of 7-day treatment regimens of levofloxacin or rifampin, alone or in combination, in rabbits with experimental prosthetic knee infections due to S. aureus 17848

Antimicrobial-Related Severe Adverse Events during Treatment of Bone and Joint Infection Due to Methicillin-Susceptible Staphylococcus aureus

Prolonged antimicrobial therapy is recommended for methicillin-susceptible Staphylococcus aureus (MSSA) bone and joint infections (BJI), but its safety profile and risk factors for severe adverse events (SAE) in clinical practice are unknown. We addressed these issues in a retrospective cohort study (2001 to 2011) analyzing antimicrobial-related SAE (defined according to the Common Terminology Criteria for Adverse Events) in 200 patients (male, 62%; median age, 60.8 years [interquartile range {IQR}, 45.5 to 74.2 years]) with MSSA BJI admitted to a reference regional center with acute (66%) or chronic arthritis (7.5%), osteomyelitis (9.5%), spondylodiscitis (16%), or orthopedic device-related infections (67%). These patients received antistaphylococcal therapy for a median of 26.6 weeks (IQR, 16.8 to 37.8 weeks). Thirty-eight SAE occurred in 30 patients (15%), with a median time delay of 34 days (IQR, 14.75 to 60.5 days), including 10 patients with hematologic reactions, 9 with cutaneomucosal reactions, 6 with acute renal injuries, 4 with hypokalemia, and 4 with cholestatic hepatitis. The most frequently implicated antimicrobials were antistaphylococcal penicillins (ASP) (13 SAE/145 patients), fluoroquinolones (12 SAE/187 patients), glycopeptides (9 SAE/101 patients), and rifampin (7 SAE/107 patients). Kaplan-Meier curves and stepwise binary logistic regression analyses were used to determine the risk factors for the occurrence of antimicrobial-related SAE. Age (odds ratio [OR], 1.479 for 10-year increase; 95% confidence interval [CI], 1.116 to 1.960; P = 0.006) appeared to be the only independent risk factor for SAE. In patients receiving ASP or rifampin, daily dose (OR, 1.028; 95% CI, 1.006 to 1.051; P = 0.014) and obesity (OR, 8.991; 95% CI, 1.453 to 55.627; P = 0.018) were associated with the occurrence of SAE. The high rate of SAE and their determinants highlighted the importance of the management and follow-up of BJI, with particular attention to be paid to older persons, especially for ASP dosage, and to rifampin dose adjustment in obese patients.     

In Vivo Effects of Cefazolin,Daptomycin, and Nafcillin in Experimental Endocarditis with a Methicillin-Susceptible Staphylococcus aureus Strain Showing an Inoculum Effect against Cefazolin

Several reports have implicated the inoculum effect that some strains of type A beta-lactamase (Bla)-producing, methicillin-susceptible Staphylococcus aureus (MSSA) show against cefazolin as the cause for clinical failures in certain serious deep-seated infections. Here, using a previously reported MSSA strain displaying this phenotype (TX0117), we obtained a Bla-cured derivative (TX0117c) with a combination of novobiocin and high temperature. Both isolates were then used in a rat endocarditis model and treated with cefazolin, nafcillin, and daptomycin, given to simulate human dosing. Animals were treated for 3 days and either sacrificed at 24 h after the last antibiotic dose (standard group) or left untreated for an additional 3 days (relapse group). With TX0117 in the standard treatment group, daptomycin and nafcillin were both significantly better than cefazolin in reducing CFU/g of vegetations, achieving mean log₁₀ reductions compared to levels in untreated rats of 7.1, 5.3, and 1.8, respectively (cefazolin versus daptomycin, P < 0.0001; cefazolin versus nafcillin, P = 0.005; daptomycin versus nafcillin, P = 0.053). In addition, cefazolin was significantly more effective in reducing vegetation titers of TX0117c than of TX0117 (mean log₁₀ reduction of 1.4 versus 5.5, respectively; P = 0.0001). Similar results were observed with animals in the relapse group. Thus, these data show that there can be an in vivo consequence of the in vitro inoculum effect that some MSSA strains display against cefazolin and indicate a specific role for Bla production using a Bla-cured derivative strain against which cefazolin regained both in vitro and in vivo activity.     

Dynamics of Clarithromycin and Azithromycin Efficacies against Experimental Haemophilus influenzae Pulmonary Infection

The dynamics of clarithromycin and azithromycin efficacy against pulmonary Haemophilus influenzae infection in rats were evaluated. Efficacy was measured by reduction in pulmonary H. influenzae burden on days 3 and 7 postinoculation. Clarithromycin therapy was effective on day 3 or 7 of therapy, while azithromycin was effective on day 7 but not on day 3 of therapy. Both macrolides produced marked efficacy against all six strains of H. influenzae tested, including four strains for which MICs were above the susceptible breakpoint (8 μg/ml) concentration of clarithromycin. The two macrolides demonstrated markedly different pharmacokinetic characteristics, with clarithromycin present in both blood and tissue, while azithromycin was concentrated primarily in tissue. During pulmonary infection in rats, H. influenzae was found in both intracellular locations and an extracellular location in the lung. Blood concentrations of clarithromycin and azithromycin approximated human pharmacokinetics, and the blood concentrations for either macrolide rarely exceeded MICs for H. influenzae. At dosages producing blood concentrations similar to values achieved clinically, clarithromycin produced efficacy on day 3 of therapy, while both clarithromycin and azithromycin were equally effective on day 7. The different dynamics of clarithromycin and azithromycin suggest that length of therapy should be considered as a key parameter in evaluations of drug efficacy.     

Efficacy of Trovafloxacin against Experimental Staphylococcus aureus Endocarditis

Trovafloxacin is a new fluoronaphthyridone chemically and functionally related to members of the fluoroquinolone class of antimicrobial agents. The in vivo efficacy of the drug was compared with that of vancomycin by using the rabbit model of left-sided endocarditis. Rabbits infected with either a nafcillin-susceptible or -resistant test strain were treated with trovafloxacin (13.3 mg/kg of body weight every 12 h) or vancomycin (25 mg/kg of body weight every 8 h) for 4 days. In comparison with untreated controls, both antimicrobial agents effectively cleared bacteremia and significantly reduced bacterial counts in vegetations and tissues of animals infected with either test strain. No resistance to trovafloxacin emerged in test strains during therapy. We conclude that in this model trovafloxacin is as efficacious as vancomycin is and may serve as a viable alternative to vancomycin for use in humans with similar infections.Trovafloxacin (CP-99,219-27) is a fluoronaphthyridone antimicrobial agent related to fluoroquinolones such as ciprofloxacin and norfloxacin (2). It has potent activity against a broad range of bacterial species, including Staphylococcus aureus (4, 5, 15, 17, 22). The antibacterial activity of trovafloxacin versus S. aureus surpasses that of ciprofloxacin, and trovafloxacin may maintain clinically relevant activity against ciprofloxacin-resistant strains (1).The in vivo activity of trovafloxacin against selected pathogens has been assessed by using animal models, and the drug has shown therapeutic efficacy comparable to that of standard forms of therapy (3, 6, 13, 21). However, the efficacy of trovafloxacin in treating serious S. aureus infections has not been determined. In order to address this issue, we compared the therapeutic activities of trovafloxacin and vancomycin by using the rabbit model of left-sided S. aureus endocarditis. This model affords a severe test of antimicrobial activity in a serious systemic infection.     

Examination of Methicillin-Resistant and Methicillin-Susceptible Staphylococcus aureus Mutants with Low-Level Fluoroquinolone Resistance

For Staphylococcus aureus, stepwise mutations result in high-level quinolone resistance. Methicillin-resistant and -susceptible quinolone-resistant, first-step mutants generated in vitro were obtained and found to be no different than those recovered from murine abscesses. Approximately 10% of all first-step mutants were resistant to ethidium bromide, and selected strains had mutations that mapped to flqB. NorA-mediated resistance among first-step mutants may be more prevalent than previously reported.     

Lack of Bactericidal Antagonism or Synergism In Vitro between Oxacillin and Vancomycin against Methicillin-Susceptible Strains of Staphylococcus aureus

With the current high prevalence of infection caused by methicillin-resistant Staphylococcus aureus (MRSA) strains but in light of the general belief that β-lactam antibiotics are more effective than vancomycin against infections caused by methicillin-susceptible S. aureus (MSSA) isolates, clinicians may utilize antistaphylococcal penicillins in combination with vancomycin for the empirical treatment of S. aureus infections. Vancomycin is considered to kill MSSA more slowly than oxacillin. Thus, we sought to evaluate the interaction of the combination of oxacillin and vancomycin on bacterial killing in vitro. Ten clinical isolates of MSSA isolated in the year 2000 were investigated. The killing observed at 24 h by vancomycin at 20 μg/ml, oxacillin at 16 μg/ml, or the combination did not differ (approximately 2.5 to 3.5 log₁₀ CFU/ml). In a separate experiment, we assessed bacterial killing in a dynamic model simulating the free plasma concentration profiles expected following the administration of a combination of vancomycin at 1 g every 12 h and oxacillin at 1 g every 6 h. The time-kill profiles of these regimens against S. aureus ATCC 29213 were comparable to those observed in the fixed-concentration experiments. Using these methods, we found no evidence that vancomycin antagonized the bactericidal effect of oxacillin or that there was any benefit from use of the combination.In many clinical settings, methicillin-resistant Staphylococcus aureus (MRSA) strains comprise such a high proportion of the S. aureus isolates that empirical treatment with vancomycin is begun when a staphylococcal infection is suspected (10, 18). However, several studies have reported that for infections due to methicillin-susceptible S. aureus (MSSA) strains, treatment with β-lactam antibiotics results in clinical outcomes better than those achieved with vancomycin (15, 18). For this reason, some clinicians use vancomycin and a β-lactam together as empirical treatment, both to provide adequate coverage for MRSA and in the belief that the β-lactam would provide superior antimicrobial activity should the organism prove to be methicillin susceptible.Small and Chambers (17) reported that vancomycin killed MSSA more slowly than nafcillin when they used time-kill methods. This observation suggests that when the antibiotics are used together, the more slowly acting bactericidal agent, vancomycin, may have the potential to antagonize the more rapid bactericidal effect of oxacillin, negating the anticipated benefit of the β-lactam. Such in vitro antagonism has been demonstrated when ß-lactams are combined with bacteriostatic agents such as erythromycin (8). In the present study, we sought to evaluate the interactions between vancomycin and oxacillin against clinical isolates of MSSA.(Part of the data presented here were presented at the 49th Interscience Conference on Antimicrobial Agents and Chemotherapy, San Francisco, CA, 12 to 15 September 2009 [abstr. E-1452].)     

Experimental phage therapy against Staphylococcus aureus in mice

The present study describes a bacteriophage (M(Sa)) active against Staphylococcus aureus, including methicillin-resistant staphylococcal strains. When inoculated into mice simultaneously with S. aureus A170 (10(8) CFU/mouse), phage (10(9) PFU) rescued 97% of the mice; when applied to nonlethal (5 x 10(6) CFU/mouse) 10-day infections, the phage also fully cleared the bacteria. The phage M(Sa), delivered inside macrophages by S. aureus, kills the intracellular staphylococci in vivo and in vitro. The phage can also prevent abscess formation and reduce the bacterial load and weight of abscesses. These results suggest a potential use of the phage for the control of both local and systemic human S. aureus infections.     

Comparative pharmacodynamics of gentamicin against Staphylococcus aureus and Pseudomonas aeruginosa

Aminoglycosides are often used to treat severe infections with gram-positive organisms. Previous studies have shown concentration-dependent killing by aminoglycosides of gram-negative bacteria, but limited data are available for gram-positive bacteria. We compared the in vitro pharmacodynamics of gentamicin against Staphylococcus aureus and Pseudomonas aeruginosa. Five S. aureus strains were examined (ATCC 29213 and four clinical isolates). Time-kill studies (TKS) in duplicate (baseline inocula of 10(7) CFU/ml) were conducted to evaluate bacterial killing in relation to increasing gentamicin concentrations (0 to 16 times the MIC). Serial samples were obtained over 24 h to quantify bacterial burden. Similar TKS with P. aeruginosa ATCC 27853 were conducted, and the time courses of the all bacterial strains were mathematically modeled for quantitative comparison. A dose fractionation study (using identical daily doses of gentamicin) in an in vitro hollow-fiber infection model (HFIM) over 5 days was subsequently used for data validation for the two ATCC strains. Model fits to the data were satisfactory; r(2) values for the S. aureus and P. aeruginosa ATCC strains were 0.915 and 0.956, respectively. Gentamicin was found to have a partially concentration-dependent killing effect against S. aureus; concentrations beyond four to 8 times the MIC did not result in significantly faster bacterial killing. In contrast, a concentration-dependent profile was demonstrated in suppressing P. aeruginosa regrowth after initial decline in bacterial burden. In HFIM, thrice-daily gentamicin dosing appeared to be superior to once-daily dosing for S. aureus, but they were similar for P. aeruginosa. Different killing profiles of gentamicin were demonstrated against S. aureus and P. aeruginosa. These results may guide optimal dosing strategies of gentamicin in S. aureus infections and warrant further investigations.     

Topoisomerase Mutations in Fluoroquinolone-Resistant and Methicillin-Susceptible and -Resistant Clinical Isolates of Staphylococcus aureus

The incidence of the various mutations in the genes encoding topoisomerase IV and DNA gyrase in fluoroquinolone-resistant clinical isolates of Staphylococcus aureus is not known. Using restriction fragment length polymorphism analysis and DNA sequencing, we found that in fluoroquinolone- and methicillin-resistant strains, mutations in grlA and gyrA are quite likely to be present together. For fluoroquinolone-resistant but methicillin-susceptible strains, mutations in grlA alone are more common.     

Pharmacodynamic Profile of Tigecycline against Methicillin-Resistant Staphylococcus aureus in an Experimental Pneumonia Model

Tigecycline (TGC) is an extended-spectrum antibiotic with activity against Staphylococcus aureus, including methicillin (meticillin)-resistant S. aureus strains, which are well-recognized pathogens in nosocomial pneumonia. The objective of this study was to characterize the exposure-response relationship for TGC against S. aureus in an immunocompromised BALB/c murine pneumonia model. Six S. aureus isolates were studied, and the TGC MICs for those isolates ranged from 0.125 to 0.5 mg/liter. The pharmacokinetics (PK) of TGC in serum and bronchoalveolar lavage (BAL) fluid were evaluated, as was the level of protein binding of the compound in this murine species. Administration of TGC at 1.56 to 150 mg/kg of body weight/day in single or two to three divided doses was used in the efficacy studies. TGC displayed linear PK and had a mean half-life of 10.9 ± 2.5 h. Efficacy was highly correlated with the area under the free concentration-time curve (fAUC)/MIC (r² = 0.93). The 80% and 50% effective exposure indexes and the stasis exposure index were similar between the isolates (means ± standard deviations, 3.04 ± 1.12, 1.84 ± 1.3, and 1.9 ± 1.5, respectively). Maximal efficacy was predicted at a 2.85-log₁₀-CFU reduction. TGC appeared to accumulate in the interstitial space, as the ratios of the fAUC from 0 to 8 h of epithelial lining fluid to plasma were 7.02, 15.11, and 23.95 for doses of 12.5, 25, and 50 mg/kg, respectively. TGC was highly effective in this murine pneumonia model. In light of current MIC distributions, the fAUC/MIC targets that we defined against S. aureus are readily achievable in humans given conventional doses of TGC.Staphylococcus aureus has long been recognized as an important cause of infection, and the emergence of S. aureus strains with the methicillin (meticillin)-resistant phenotype (methicillin-resistant S. aureus [MRSA]) has further complicated management. Both community-acquired MRSA (CA-MRSA) and hospital-acquired MRSA (HA-MRSA) strains have been associated with severe and difficult-to-treat infections. While the most common site of staphylococcal infection is the skin and skin structures, the surveillance of 8,792 invasive MRSA cases in the United States showed that pneumonia is the second most common clinical manifestation of MRSA infection (13.3% overall; 14% of the strains were CA-MRSA and 28% were HA-MRSA) (9).Tigecycline (TGC) is a broad-spectrum glycylcycline with efficacy against gram-positive and gram-negative bacteria, including drug-resistant bacteria such as MRSA. The MIC₉₀ of TGC against methicillin-susceptible S. aureus (MSSA) and MRSA strains is reported to be ≤0.25 mg/liter (6). TGC is approved by the FDA for use for the treatment of complicated skin and skin structure infections and complicated intra-abdominal infections. Pneumonia is an important clinical manifestation of infection with drug-resistant bacteria; therefore, many in vivo and in vitro studies of TGC for the treatment of lower respiratory infection are ongoing (data available at http://www.clinicaltrialssearch.org/tigecycline_versus_imipenemcilastatin_for_the_treatment_of_subjects_with_nosocomial_pneumonia.html and http://www.medicalnewstoday.com/articles/53035.php).Previously, we demonstrated the efficacy of TGC against Acinetobacter spp. in a murine pneumonia model (10) and against S. aureus in a murine thigh infection model (3). We also found that TGC penetrated well into lung tissue, as displayed by high concentrations in bronchoalveolar (BAL) fluid (4). In the present study, we aimed to explore the exposure-response relationship for TGC against S. aureus in an immunocompromised BALB/c murine pneumonia model.     

Pharmacodynamics of Ceftaroline against Staphylococcus aureus Studied in an In Vitro Pharmacokinetic Model of Infection

An in vitro single-compartment dilutional pharmacokinetic model was used to study the pharmacodynamics of ceftaroline against Staphylococcus aureus (both methicillin-susceptible S. aureus [MSSA] and methicillin-resistant S. aureus [MRSA]). Mean serum free concentrations of ceftaroline (the active metabolite of the prodrug ceftaroline fosamil) dosed in humans at 600 mg every 12 h (q12h) were simulated, and activities against 12 S. aureus strains (3 MSSA strains and 9 MRSA strains, 3 of which had a vancomycin-intermediate phenotype) were determined. Ceftaroline produced 2.5- to 4.0-log₁₀-unit reductions in viable counts by 24 h with all strains and a 0.5- to 4.0-log-unit drop in counts at 96 h. The antibacterial effect could not be related to the strain MIC across the ceftaroline MIC range from 0.12 to 2.0 μg/ml. In dose-ranging studies, the cumulative percentage of a 24-h period that the free drug concentration exceeded the MIC under steady-state pharmacokinetic conditions (fT_MIC) of 24.5% ± 8.9% was associated with a 24-h bacteriostatic effect, one of 27.8% ± 9.5% was associated with a −1-log-unit drop, and one of 32.1% ± 8.1% was associated with a −2-log-unit drop. The MSSA and MRSA strains had similar fT_MIC values. fT_MIC values increased with increasing duration of exposure up to 96 h. Changes in ceftaroline population analysis profiles were related to fT_MIC. fT_MICs of <50% were associated with growth on 4× MIC recovery plates at 96 h of drug exposure. These data support the use of ceftaroline fosamil at doses of 600 mg q12h to treat S. aureus strains with MICs of ≤2 μg/ml. An fT_MIC of 25 to 30% would make a suitable pharmacodynamic index target, but fT_MIC values of ≥50% are needed to suppress the emergence of resistance and require clinical evaluation.