Efficacy of Fosfomycin Compared to Vancomycin in Treatment of Implant-Associated Chronic Methicillin-Resistant Staphylococcus aureus Osteomyelitis in Rats

Fosfomycin monotherapy was compared to therapy with vancomycin for the treatment of implant-associated methicillin-resistant Staphylococcus aureus (MRSA) osteomyelitis in an experimental rat model. The proximal tibiae were inoculated with 15 μl of a suspension containing 1 × 10⁸ to 5 × 10⁸ CFU/ml of a clinical isolate of MRSA with simultaneous insertion of a titanium wire. Four weeks later, treatment was started for 28 days with either 50 mg/kg of body weight vancomycin intraperitoneally twice daily (n = 11) or 75 mg/kg fosfomycin intraperitoneally once daily (n = 10). Eleven animals were left untreated. After treatment, quantitative cultures from bone were found to be positive for MRSA in all animals in the untreated group (median, 3.29 × 10⁶ CFU/g of bone) and the vancomycin group (median, 3.03 × 10⁵ CFU/g of bone). In the fosfomycin group, MRSA was detectable in 2 out of 10 (20%) animals (3.42 × 10² and 1.51 × 10³ CFU/g of bone). Vancomycin was superior to the no-drug control (P = 0.002), and fosfomycin was superior to the no-drug control and vancomycin (P < 0.001). The cultures from the wires were positive in all untreated animals (median, 2.5 × 10³ CFU/implant), in 10 animals in the vancomycin group (median, 1.15 × 10³ CFU/implant), and negative in all animals in the fosfomycin group. Based on the bacterial counts from the implants, vancomycin was not superior to the no-drug control (P = 0.324), and fosfomycin was superior to the no-drug control and vancomycin (P < 0.001). No emergence of resistance was observed. In conclusion, it was demonstrated that fosfomycin monotherapy is highly effective for the treatment of experimental implant-associated MRSA osteomyelitis.     

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.     

A Novel Approach Utilizing Biofilm Time-Kill Curves To Assess the Bactericidal Activity of Ceftaroline Combinations against Biofilm-Producing Methicillin-Resistant Staphylococcus aureus

Medical device infections frequently require combination therapy. Beta-lactams combined with glycopeptides/lipopeptides are bactericidal against methicillin-resistant Staphylococcus aureus (MRSA). Novel macrowell kill-curve methods tested synergy between ceftaroline or cefazolin plus daptomycin, vancomycin, or rifampin against biofilm-producing MRSA. Ceftaroline combinations demonstrated the most pronounced bacterial reductions. Ceftaroline demonstrated greatest kill with daptomycin (4.02 ± 0.59 log₁₀ CFU/cm²), compared to combination with vancomycin (3.36 ± 0.35 log₁₀ CFU/cm²) or rifampin (2.68 ± 0.61 log₁₀ CFU/cm²). These data suggest that beta-lactam combinations are useful against MRSA biofilms.     

Daptomycin, fosfomycin, or both for treatment of methicillin-resistant Staphylococcus aureus osteomyelitis in an experimental rat model

The in vivo activities of daptomycin, fosfomycin, and a combination of both antibiotics against a clinical isolate of methicillin-resistant Staphylococcus aureus (daptomycin MIC, 0.25 μg/ml; fosfomycin MIC, 0.5 μg/ml) were evaluated in a rat model of osteomyelitis. A total of 37 rats with experimental osteomyelitis were treated for 4 weeks with either 60 mg/kg of body weight of daptomycin subcutaneously once daily, 75 mg/kg fosfomycin intraperitoneally once daily, a combination of both drugs, or a saline placebo. After the completion of treatment, animals were euthanized, and the infected tibiae were processed for quantitative bacterial culture. Bone cultures were found to be positive for methicillin-resistant S. aureus in 9 of 9 (100%) animals of the placebo group, in 9 of 9 (100%) animals treated with daptomycin, in 1 of 10 (10%) fosfomycin-treated rats, and in 1 of 9 (22.2%) rats comprising the combination group. Results of bacterial counts in the bone samples were expressed as log(10) CFU/g of bone and analyzed by using the Mann-Whitney U test followed by Bonferroni's multiple-comparison test. Based on bacterial counts, treatment with daptomycin was significantly superior to placebo, although it remained inferior to treatment with fosfomycin. No synergistic or antagonistic effect was observed for the combination therapy. No development of resistance against daptomycin or fosfomycin was observed after the 4-week treatment period.     

In Vitro Pharmacodynamics of Human Simulated Exposures of Ceftaroline and Daptomycin against MRSA,hVISA, and VISA with and without Prior Vancomycin Exposure

The effects of prior vancomycin exposure on ceftaroline and daptomycin therapy against methicillin-resistant Staphylococcus aureus (MRSA) have not been widely studied. Humanized free-drug exposures of vancomycin at 1 g every 12 h (q12h), ceftaroline at 600 mg q12h, and daptomycin at 10 mg/kg of body weight q24h were simulated in a 96-h in vitro pharmacodynamic model against three MRSA isolates, including one heteroresistant vancomycin-intermediate S. aureus (hVISA) isolate and one VISA isolate. A total of five regimens were tested: vancomycin, ceftaroline, and daptomycin alone for the entire 96 h, and then sequential therapy with vancomycin for 48 h followed by ceftaroline or daptomycin for 48 h. Microbiological responses were measured by the changes in log₁₀ CFU during 96 h from baseline. Control isolates grew to 9.16 ± 0.32, 9.13 ± 0.14, and 8.69 ± 0.28 log₁₀ CFU for MRSA, hVISA, and VISA, respectively. Vancomycin initially achieved ≥3 log₁₀ CFU reductions against the MRSA and hVISA isolates, followed by regrowth beginning at 48 h; minimal activity was observed against VISA. The change in 96-h log₁₀ CFU was largest for sequential therapy with vancomycin followed by ceftaroline (−5.22 ± 1.2, P = 0.010 versus ceftaroline) and for sequential therapy with vancomycin followed by ceftaroline (−3.60 ± 0.6, P = 0.037 versus daptomycin), compared with daptomycin (−2.24 ± 1.0), vancomycin (−1.40 ± 1.8), and sequential therapy with vancomycin followed by daptomycin (−1.32 ± 1.0, P > 0.5 for the last three regimens). Prior exposure of vancomycin at 1 g q12h reduced the initial microbiological response of daptomycin, particularly for hVISA and VISA isolates, but did not affect the response of ceftaroline. In the scenario of poor vancomycin response for high-inoculum MRSA infection, a ceftaroline-containing regimen may be preferred.     

Activities of Fosfomycin and Rifampin on Planktonic and Adherent Enterococcus faecalis Strains in an Experimental Foreign-Body Infection Model

Enterococcal implant-associated infections are difficult to treat because antibiotics generally lack activity against enterococcal biofilms. We investigated fosfomycin, rifampin, and their combinations against planktonic and adherent Enterococcus faecalis (ATCC 19433) in vitro and in a foreign-body infection model. The MIC/MBC_log values were 32/>512 μg/ml for fosfomycin, 4/>64 μg/ml for rifampin, 1/2 μg/ml for ampicillin, 2/>256 μg/ml for linezolid, 16/32 μg/ml for gentamicin, 1/>64 μg/ml for vancomycin, and 1/5 μg/ml for daptomycin. In time-kill studies, fosfomycin was bactericidal at 8× and 16× MIC, but regrowth of resistant strains occurred after 24 h. With the exception of gentamicin, no complete inhibition of growth-related heat production was observed with other antimicrobials on early (3 h) or mature (24 h) biofilms. In the animal model, fosfomycin alone or in combination with daptomycin reduced planktonic counts by ≈4 log₁₀ CFU/ml below the levels before treatment. Fosfomycin cleared planktonic bacteria from 74% of cage fluids (i.e., no growth in aspirated fluid) and eradicated biofilm bacteria from 43% of cages (i.e., no growth from removed cages). In combination with gentamicin, fosfomycin cleared 77% and cured 58% of cages; in combination with vancomycin, fosfomycin cleared 33% and cured 18% of cages; in combination with daptomycin, fosfomycin cleared 75% and cured 17% of cages. Rifampin showed no activity on planktonic or adherent E. faecalis, whereas in combination with daptomycin it cured 17% and with fosfomycin it cured 25% of cages. Emergence of fosfomycin resistance was not observed in vivo. In conclusion, fosfomycin showed activity against planktonic and adherent E. faecalis. Its role against enterococcal biofilms should be further investigated, especially in combination with rifampin and/or daptomycin treatment.     

Efficacy of Daptomycin in Implant-Associated Infection Due to Methicillin-Resistant Staphylococcus aureus: Importance of Combination with Rifampin

Limited treatment options are available for implant-associated infections caused by methicillin (meticillin)-resistant Staphylococcus aureus (MRSA). We compared the activity of daptomycin (alone and with rifampin [rifampicin]) with the activities of other antimicrobial regimens against MRSA ATCC 43300 in the guinea pig foreign-body infection model. The daptomycin MIC and the minimum bactericidal concentration in logarithmic phase and stationary growth phase of MRSA were 0.625, 0.625, and 20 μg/ml, respectively. In time-kill studies, daptomycin showed rapid and concentration-dependent killing of MRSA in stationary growth phase. At concentrations above 20 μg/ml, daptomycin reduced the counts by >3 log₁₀ CFU/ml in 2 to 4 h. In sterile cage fluid, daptomycin peak concentrations of 23.1, 46.3, and 53.7 μg/ml were reached 4 to 6 h after the administration of single intraperitoneal doses of 20, 30, and 40 mg/kg of body weight, respectively. In treatment studies, daptomycin alone reduced the planktonic MRSA counts by 0.3 log₁₀ CFU/ml, whereas in combination with rifampin, a reduction in the counts of >6 log₁₀ CFU/ml was observed. Vancomycin and daptomycin (at both doses) were unable to cure any cage-associated infection when they were given as monotherapy, whereas rifampin alone cured the infections in 33% of the cages. In combination with rifampin, daptomycin showed cure rates of 25% (at 20 mg/kg) and 67% (at 30 mg/kg), vancomycin showed a cure rate of 8%, linezolid showed a cure rate of 0%, and levofloxacin showed a cure rate of 58%. In addition, daptomycin at a high dose (30 mg/kg) completely prevented the emergence of rifampin resistance in planktonic and adherent MRSA cells. Daptomycin at a high dose, corresponding to 6 mg/kg in humans, in combination with rifampin showed the highest activity against planktonic and adherent MRSA. Daptomycin plus rifampin is a promising treatment option for implant-associated MRSA infections.Implants are increasingly used in modern medicine to replace a compromised biological function or missing anatomical structure. Periprosthetic infections represent a devastating complication, causing high rates of morbidity and consuming considerable health care resources. Implant-associated infections are caused by microorganisms growing adherent to the device surface and embedded in an extracellular polymeric matrix, a complex three-dimensional structure called a microbial biofilm (8). Bacterial communities in biofilms cause persistent infection due to increased resistance to antibiotics and the immune system and the difficulty with eradicating them from the implant (6).Staphylococcus aureus is one of the leading pathogens causing implant-associated infections. Successful treatment requires the use of bactericidal drugs acting on surface-adhering microorganisms, which predominantly exist in the stationary growth phase. Previous in vitro, experimental, and clinical studies demonstrated that rifampin (rifampicin)-containing antimicrobial regimens were able to eradicate staphylococcal biofilms and cure implant-associated infections (23, 25). Quinolones are often used in combination with rifampin in order to prevent the emergence of rifampin resistance (4, 19, 21). However, methicillin (meticillin)-resistant S. aureus (MRSA) strains are often resistant to quinolones. In addition, MRSA strains were recently shown to have decreased susceptibility to vancomycin, reducing the efficacy of this drug. Therefore, alternative drugs for use in combination with rifampin against implant-associated infections are needed (12, 20).Daptomycin is a negatively charged cyclic lipopeptide with bactericidal activity against gram-positive organisms, including MRSA (17). The drug inserts into the bacterial cytoplasmic membrane in a calcium-dependent fashion, leading to rapid cell death without lysis, and causing only minimal inflammation (15). Daptomycin has been well tolerated in healthy volunteers dosed with up to 12 mg/kg of body weight intravenously for 14 days (2). Only limited data on the use of daptomycin in combination with rifampin against staphylococcal implant-associated infections are available.In this study, we investigated the activity of daptomycin against MRSA ATCC 43300 in vitro. In addition, we evaluated the activity of daptomycin in combination with rifampin in a cage-associated infection model in guinea pigs and compared the efficacy of the treatment with the efficacies of three other antibiotics commonly used against MRSA, vancomycin, linezolid, and levofloxacin (alone and in combination with rifampin).(Part of the results of the present study were presented at the 48th Interscience Conference on Antimicrobial Agents and Chemotherapy, Washington, DC, 24 to 29 October 2008 [abstr. B-1000].)     

Efficacy of Teicoplanin and Autoradiographic Diffusion Pattern of [14C]Teicoplanin in Experimental Staphylococcus aureus Infection of Joint Prostheses

Prosthesis infections are difficult to cure. Infection with methicillin-resistant staphylococci is becoming more common in patients with orthopedic implants. Using a recently developed model of methicillin-resistant Staphylococcus aureus (MRSA) infection of a knee prosthesis, we compared the efficacies of teicoplanin and vancomycin. [¹⁴C]teicoplanin diffusion in this model was also studied by autoradiography. A partial knee replacement was performed with a silicone implant fitting into the intramedullary canal of the tibia, and 10⁷ CFU of MRSA was injected into the knee. Treatment with teicoplanin or vancomycin (20 or 60 mg/kg of body weight, respectively, given intramuscularly twice daily) was started 7 days after inoculation and was continued for 7 days. The teicoplanin and vancomycin MICs for MRSA were 1 μg/ml. Mean peak and trough levels in serum were 39.1 and 23.5 μg/ml, respectively, for teicoplanin and 34.4 and 18.5 μg/ml, respectively, for vancomycin. Fifteen days after the end of therapy, the animals were killed and their tibias were removed, pulverized, and quantitatively cultured. Teicoplanin and vancomycin significantly reduced (P < 0.05) the bacterial density (2.7 ± 1.3 and 3.3 ± 1.6 log₁₀ CFU/g of bone, respectively) compared to those for the controls (5.04 ± 1.4 log₁₀ CFU/g of bone). The bacterial covents of teicoplanin- and vancomycin-treated rabbits were comparable. The [¹⁴C]teicoplanin autoradiographic diffusion patterns in rabbits with prostheses, two of which were uninfected and two of which were infected, were studied 15 days after inoculation. Sixty minutes after the end of an infusion of 250 μCi of [¹⁴C]teicoplanin, autoradiography showed that in the infected animals, the highest levels of radioactivity were located around the prosthesis and in the periosteum, bone marrow, and trabecular bone. Radioactivity was less intense in epiphyseal disk cartilage, femoral cartilage, articular ligaments, and muscles and was weak in compact bone. A similar distribution pattern was seen in uninfected rabbits. Thus, teicoplanin may represent an effective alternative therapy for the treatment of these infections.     

Evaluation of the Novel Combination of High-Dose Daptomycin plus Trimethoprim-Sulfamethoxazole against Daptomycin-Nonsusceptible Methicillin-Resistant Staphylococcus aureus Using an In Vitro Pharmacokinetic/Pharmacodynamic Model of Simulated Endocardial Vegetations

Daptomycin-nonsusceptible (DNS) Staphylococcus aureus is found in difficult-to-treat infections, and the optimal therapy is unknown. We investigated the activity of high-dose (HD) daptomycin plus trimethoprim-sulfamethoxazole de-escalated to HD daptomycin or trimethoprim-sulfamethoxazole against 4 clinical DNS methicillin-resistant S. aureus (MRSA) isolates in an in vitro pharmacokinetic/pharmacodynamic model of simulated endocardial vegetations (10⁹ CFU/g). Simulated regimens included HD daptomycin at 10 mg/kg/day for 14 days, trimethoprim-sulfamethoxazole at 160/800 mg every 12 h for 14 days, HD daptomycin plus trimethoprim-sulfamethoxazole for 14 days, and the combination for 7 days de-escalated to HD daptomycin for 7 days and de-escalated to trimethoprim-sulfamethoxazole for 7 days. Differences in CFU/g (at 168 and 336 h) were evaluated by analysis of variance (ANOVA) with a Tukey''s post hoc test. Daptomycin MICs were 4 μg/ml (SA H9749-1, vancomycin-intermediate Staphylococcus aureus; R6212, heteroresistant vancomycin-intermediate Staphylococcus aureus) and 2 μg/ml (R5599 and R5563). Trimethoprim-sulfamethoxazole MICs were ≤0.06/1.19 μg/ml. HD daptomycin plus trimethoprim-sulfamethoxazole displayed rapid bactericidal activity against SA H9749-1 (at 7 h) and R6212 (at 6 h) and bactericidal activity against R5599 (at 72 h) and R5563 (at 36 h). A ≥8 log₁₀ CFU/g decrease was observed with HD daptomycin plus trimethoprim-sulfamethoxazole against all strains (at 48 to 144 h), which was maintained with de-escalation to HD daptomycin or trimethoprim-sulfamethoxazole at 336 h. The combination for 14 days and the combination for 7 days de-escalated to HD daptomycin or trimethoprim-sulfamethoxazole was significantly better than daptomycin monotherapy (P < 0.05) and trimethoprim-sulfamethoxazole monotherapy (P < 0.05) at 168 and 336 h. Combination therapy followed by de-escalation offers a novel bactericidal therapeutic alternative for high-inoculum, serious DNS MRSA infections.     

Evaluation of Ceftaroline,Vancomycin, Daptomycin,or Ceftaroline plus Daptomycin against Daptomycin-Nonsusceptible Methicillin-Resistant Staphylococcus aureus in an In Vitro Pharmacokinetic/Pharmacodynamic Model of Simulated Endocardial Vegetations

Infective endocarditis (IE) caused by methicillin-resistant Staphylococcus aureus (MRSA) with reduced susceptibility to vancomycin and daptomycin has few adequate therapeutic options. Ceftaroline (CPT) is bactericidal against daptomycin (DAP)-nonsusceptible (DNS) and vancomycin-intermediate MRSA, but supporting data are limited for IE. This study evaluated the activities of ceftaroline, vancomycin, daptomycin, and the combination of ceftaroline plus daptomycin against DNS MRSA in a pharmacokinetic/pharmacodynamic (PK/PD) model of simulated endocardial vegetations (SEVs). Simulations of ceftaroline-fosamil (600 mg) every 8 h (q8h) (maximum concentration of drug in serum [C_max], 21.3 mg/liter; half-life [t_1/2], 2.66 h), daptomycin (10 mg/kg of body weight/day) (C_max, 129.7 mg/liter; t_1/2, 8 h), vancomycin (1 g) q8h (minimum concentration of drug in serum [C_min], 20 mg/liter; t_1/2, 5 h), and ceftaroline plus daptomycin were evaluated against 3 clinical DNS, vancomycin-intermediate MRSA in a two-compartment, in vitro, PK/PD SEV model over 96 h with a starting inoculum of ∼8 log₁₀ CFU/g. Bactericidal activity was defined as a ≥3-log₁₀ CFU/g reduction from the starting inoculum. Therapeutic enhancement of combinations was defined as ≥2-log₁₀ CFU/g reduction over the most active agent alone. MIC values for daptomycin, vancomycin, and ceftaroline were 4 mg/liter, 4 to 8 mg/liter, and 0.5 to 1 mg/liter, respectively, for all strains. At simulated exposures, vancomycin was bacteriostatic, but daptomycin and ceftaroline were bactericidal. By 96 h, ceftaroline monotherapy offered significantly improved killing compared to other agents against one strain. The combination of DAP plus CPT demonstrated therapeutic enhancement, resulting in significantly improved killing versus either agent alone against 2/3 (67%) strains. CPT demonstrated bactericidal activity against DNS, vancomycin-intermediate MRSA at high bacterial densities. Ceftaroline plus daptomycin may offer more rapid and sustained activity against some MRSA in the setting of high-inoculum infections like IE and should also be considered.     

Fosfomycin Synergy In Vitro with Amoxicillin,Daptomycin, and Linezolid against Vancomycin-Resistant Enterococcus faecium from Renal Transplant Patients with Infected Urinary Stents

Fosfomycin is a potential option for vancomycin-resistant enterococcus (VRE) infections despite limited in vitro and clinical data. In this study, 32 VRE isolates from renal transplant patients with urinary stent infections were susceptible to fosfomycin, daptomycin, and linezolid and resistant to amoxicillin, minocycline, and nitrofurantoin based on their MIC₅₀s and MIC₉₀s. Fosfomycin was bacteriostatic at 0.5 to 16× the MIC (32 to 2,048 μg/ml); synergy occurred when fosfomycin was combined with daptomycin (2.8 to 3.9 log₁₀ CFU/ml kill; P < 0.001) or amoxicillin (2.6 to 3.4; P < 0.05). These combinations may be potent options to treat VRE urinary infections pending investigation of clinical efficacy.     

Ceftaroline-Fosamil Efficacy against Methicillin-Resistant Staphylococcus aureus in a Rabbit Prosthetic Joint Infection Model

Ceftaroline (CPT), the active metabolite of the prodrug ceftaroline-fosamil (CPT-F), demonstrates in vitro bactericidal activity against methicillin-resistant Staphylococcus aureus (MRSA) and is effective in rabbit models of difficult-to-treat MRSA endocarditis and acute osteomyelitis. However, its in vivo efficacy in a prosthetic joint infection (PJI) model is unknown. Using a MRSA-infected knee PJI model in rabbits, the efficacies of CPT-F or vancomycin (VAN) alone and combined with rifampin (RIF) were compared. After each partial knee replacement with a silicone implant that fit into the tibial intramedullary canal was performed, 5 × 10⁷ MRSA CFU (MICs of 0.38, 0.006, and 1 mg/liter for CPT, RIF, and VAN, respectively) was injected into the knee. Infected animals were randomly assigned to receive no treatment (controls) or CPT-F (60 mg/kg of body weight intramuscularly [i.m.]), VAN (60 mg/kg i.m.), CPT-F plus RIF (10 mg/kg i.m.), or VAN plus RIF starting 7 days postinoculation and lasting for 7 days. Surviving bacteria in crushed tibias were counted 3 days after ending treatment. Although the in vivo mean log₁₀ CFU/g of CPT-treated (3.0 ± 0.9, n = 12) and VAN-treated (3.5 ± 1.1, n = 12) crushed bones was significantly lower than those of controls (5.6 ± 1.1, n = 14) (P < 0.001), neither treatment fully sterilized the bones (3/12 were sterile with each treatment). The mean log₁₀ CFU/g values for the antibiotics in combination with RIF were 1.9 ± 0.5 (12/14 were sterile) for CPT-F and 1.9 ± 0.5 (12/14 were sterile) for VAN. In this MRSA PJI model, the efficacies of CPT-F and VAN did not differ; thus, CPT appears to be a promising antimicrobial agent for the treatment of MRSA PJIs.     

Activities of Fosfomycin,Tigecycline, Colistin,and Gentamicin against Extended-Spectrum-β-Lactamase-Producing Escherichia coli in a Foreign-Body Infection Model

Limited antimicrobial agents are available for the treatment of implant-associated infections caused by fluoroquinolone-resistant Gram-negative bacilli. We compared the activities of fosfomycin, tigecycline, colistin, and gentamicin (alone and in combination) against a CTX-M15-producing strain of Escherichia coli (Bj HDE-1) in vitro and in a foreign-body infection model. The MIC and the minimal bactericidal concentration in logarithmic phase (MBC_log) and stationary phase (MBC_stat) were 0.12, 0.12, and 8 μg/ml for fosfomycin, 0.25, 32, and 32 μg/ml for tigecycline, 0.25, 0.5, and 2 μg/ml for colistin, and 2, 8, and 16 μg/ml for gentamicin, respectively. In time-kill studies, colistin showed concentration-dependent activity, but regrowth occurred after 24 h. Fosfomycin demonstrated rapid bactericidal activity at the MIC, and no regrowth occurred. Synergistic activity between fosfomycin and colistin in vitro was observed, with no detectable bacterial counts after 6 h. In animal studies, fosfomycin reduced planktonic counts by 4 log₁₀ CFU/ml, whereas in combination with colistin, tigecycline, or gentamicin, it reduced counts by >6 log₁₀ CFU/ml. Fosfomycin was the only single agent which was able to eradicate E. coli biofilms (cure rate, 17% of implanted, infected cages). In combination, colistin plus tigecycline (50%) and fosfomycin plus gentamicin (42%) cured significantly more infected cages than colistin plus gentamicin (33%) or fosfomycin plus tigecycline (25%) (P < 0.05). The combination of fosfomycin plus colistin showed the highest cure rate (67%), which was significantly better than that of fosfomycin alone (P < 0.05). In conclusion, the combination of fosfomycin plus colistin is a promising treatment option for implant-associated infections caused by fluoroquinolone-resistant Gram-negative bacilli.     

Addition of Gentamicin or Rifampin Does Not Enhance the Effectiveness of Daptomycin in Treatment of Experimental Endocarditis Due to Methicillin-Resistant Staphylococcus aureus

This study evaluated the activity of daptomycin combined with either gentamicin or rifampin against three methicillin-resistant Staphylococcus aureus (MRSA) clinical isolates in vitro and one isolate in vivo against a representative strain (MRSA-572). Time-kill experiments showed that daptomycin was bactericidal against these strains at concentrations over the MIC. Daptomycin at sub-MIC concentrations plus gentamicin at 1× and 2× the MIC yielded synergy, while the addition of rifampin at 2 to 4 μg/ml resulted in indifference (two strains) or antagonism (one strain). The in vivo activity of daptomycin (6 mg/kg of body weight once a day) was evaluated ± gentamicin (1 mg/kg intravenously [i.v.] every 8 h [q8h]) or rifampin (300 mg i.v. q8h) in a rabbit model of infective endocarditis by simulating human pharmacokinetics. Daptomycin plus gentamicin (median, 0 [interquartile range, 0 to 2] log₁₀ CFU/g vegetation) was as effective as daptomycin alone (0 [0 to 2] log₁₀ CFU/g vegetation) in reducing the density of bacteria in valve vegetations (P = 0.83), and both were more effective than daptomycin plus rifampin (3 [2 to 3.5] log₁₀ CFU/g vegetation; P < 0.05) for the strain studied. In addition, daptomycin sterilized a ratio of vegetations that was similar to that of daptomycin plus gentamicin (10/15 [67%] versus 9/15 [60%]; P = 0.7), and both regimens did so more than daptomycin plus rifampin (3/15 [20%]; P = 0.01 and P = 0.02, respectively). No statistical difference was noted between daptomycin plus gentamicin and daptomycin alone for MRSA treatment. In the combination arm, all isolates from vegetations remained susceptible to daptomycin, gentamicin, and rifampin. Sixty-one percent of the isolates (8/13) acquired resistance to rifampin during monotherapy. In the daptomycin arm, resistance was detected in only one case, in which the daptomycin MIC rose to 2 μg/ml among the recovered bacteria. In conclusion, the addition of gentamicin or rifampin does not enhance the effectiveness of daptomycin in the treatment of experimental endocarditis due to MRSA.Staphylococcus aureus is a common cause of infective endocarditis (IE), with methicillin-resistant S. aureus (MRSA) strains found in up to one-third of all cases (11, 28). Due to multidrug resistance among many strains, vancomycin is the standard therapy for IE caused by MRSA (1). However, vancomycin therapy has been associated with poor outcomes that may be explained by the drug''s slow bactericidal activity and insufficient diffusion into valve vegetations (5, 10, 23).Daptomycin is a cyclic lipopeptide that is rapidly bactericidal against gram-positive pathogens such as MRSA, including strains that exhibit resistance to vancomycin. It is approved for the treatment of skin and soft tissue infections, S. aureus bacteremia, and right-sided native valve endocarditis (6). However, there is limited information regarding the efficacy of daptomycin in the treatment of left-sided native valve IE caused by MRSA. In a randomized clinical trial (10), none of the patients with left-sided endocarditis treated with daptomycin at 6 mg/kg of body weight/day were cured, and postmarketing registry data (24) revealed a successful clinical outcome in only 9 out of 15 cases (60%). Therefore, given the lack of efficacy data with daptomycin monotherapy in left-sided MRSA endocarditis, the continued evaluation of methods to enhance the activity of daptomycin is warranted. It is unknown whether daptomycin''s activity against MRSA may be improved by combining it with one or more additional antibiotics to produce a potentially additive or synergistic effect. Gentamicin has been shown to augment daptomycin''s activity against strains of MRSA in vitro (4, 20, 35). The combination of daptomycin plus rifampin has demonstrated additive activity against MRSA in vitro (4) and has enhanced activity against MRSA in vivo (4, 32). The aim of this study was to evaluate the in vitro activity of daptomycin combined with gentamicin or rifampin against MRSA and compare treatment with daptomycin alone to treatment with both combinations in experimental MRSA aortic valve endocarditis using a human-adapted pharmacokinetic model.(This work was previously presented at the 47th Interscience Conference on Antimicrobial Agents and Chemotherapy [ICAAC], Chicago, IL, 17 to 20 September 2007 [29a] and at the 48th Annual ICAAC-IDSA Annual Meeting, Washington, DC, 25 to 28 October 2008 [29b].)     

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.     

In vitro synergism between daptomycin and fosfomycin against Enterococcus faecalis isolates with high-level gentamicin resistance.

Daptomycin and fosfomycin are two agents which inhibit different steps in peptidoglycan synthesis. We studied the in vitro activities of these drugs, alone and in combination, by time-kill techniques against 21 clinical isolates of Enterococcus (Streptococcus) faecalis demonstrating high-level resistance to gentamicin. Combinations of fosfomycin and daptomycin exhibited synergistic bactericidal activity (100-fold decrease in CFU per milliliter at 24 h compared with daptomycin alone) against all strains (mean +/- standard deviation of increment in killing = 2.7 +/- 0.7 log10 CFU/ml). In a subgroup of strains against which daptomycin (5 micrograms/ml) alone was bactericidal (greater than 3 log10 killing), synergistic activity was demonstrable only when the concentration of daptomycin was lowered to 0.25 to 0.5 microgram/ml. A 50% dilution of human serum diminished the bactericidal activity of daptomycin alone at 24 h but did not affect killing observed with the daptomycin-fosfomycin combination. The inhibition of peptidoglycan synthesis by the combination was greater than the inhibition observed with either drug alone. The combination of daptomycin and fosfomycin exhibited consistent synergistic bactericidal activity against strains of E. faecalis possessing high-level resistance to gentamicin. This synergism may be the result of sequential inhibition of early steps in peptidoglycan synthesis.     

Characterization of High-Level Daptomycin Resistance in Viridans Group Streptococci Developed upon In Vitro Exposure to Daptomycin

Viridans group streptococci (VGS) are part of the normal flora that may cause bacteremia, often leading to endocarditis. We evaluated daptomycin against four clinical strains of VGS (MICs = 1 or 2 μg/ml) using an in vitro-simulated endocardial vegetation model, a simulated bacteremia model, and kill curves. Daptomycin exposure was simulated at 6 mg/kg of body weight and 8 mg/kg every 24 h for endocardial and bacteremia models. Total drug concentrations were used for analyses containing protein (albumin and pooled human serum), and free (unbound) drug concentrations (93% protein bound) were used for analyses not containing protein. Daptomycin MICs in the presence of protein were significantly higher than those in the absence of protein. Despite MICs below or at the susceptible breakpoint, all daptomycin regimens demonstrated limited kill in both pharmacodynamic models. A reduction of approximately 1 to 2 log₁₀ CFU was seen for all isolates and dosages except daptomycin at 6 mg/kg, which achieved a reduction of 2.7 log₁₀ CFU/g against one strain (Streptococcus gordonii 1649) in the endocardial model. Activity was similar in both pharmacodynamic models in the presence or absence of protein. Similar activity was noted in the kill curves over all multiples of the MIC. Regrowth by 24 h was seen even at 8× MIC. Postexposure daptomycin MICs for both pharmacodynamic models increased to >256 μg/ml for all isolates by 24 and 72 h. Despite susceptibility to daptomycin by standard MIC methods, these VGS developed high-level daptomycin resistance (HLDR) after a short duration following drug exposure not attributed to modification or inactivation of daptomycin. Further evaluation is warranted to determine the mechanism of resistance and clinical implications.     

Antimicrobial Activity against Intraosteoblastic Staphylococcus aureus

Although Staphylococcus aureus persistence in osteoblasts, partly as small-colony variants (SCVs), can contribute to bone and joint infection (BJI) relapses, the intracellular activity of antimicrobials is not currently considered in the choice of treatment strategies for BJI. Here, antistaphylococcal antimicrobials were evaluated for their intraosteoblastic activity and their impact on the intracellular emergence of SCVs in an ex vivo osteoblast infection model. Osteoblastic MG63 cells were infected for 2 h with HG001 S. aureus. After killing the remaining extracellular bacteria with lysostaphin, infected cells were incubated for 24 h with antimicrobials at the intraosseous concentrations reached with standard therapeutic doses. Intracellular bacteria and SCVs were then quantified by plating cell lysates. A bactericidal effect was observed with fosfomycin, linezolid, tigecycline, oxacillin, rifampin, ofloxacin, and clindamycin, with reductions in the intracellular inocula of −2.5, −3.1, −3.9, −4.2, −4.9, −4.9, and −5.2 log₁₀ CFU/100,000 cells, respectively (P < 10⁻⁴). Conversely, a bacteriostatic effect was observed with ceftaroline and teicoplanin, whereas vancomycin and daptomycin had no significant impact on intracellular bacterial growth. Ofloxacin, daptomycin, and vancomycin significantly limited intracellular SCV emergence. Overall, ofloxacin was the only molecule to combine an excellent intracellular activity while limiting the emergence of SCVs. These data provide a basis for refining the choice of antibiotics to prioritise in the management of BJI, justifying the combination of a fluoroquinolone for its intracellular activity with an anti-biofilm molecule, such as rifampin.     

Ceftaroline Increases Membrane Binding and Enhances the Activity of Daptomycin against Daptomycin-Nonsusceptible Vancomycin-Intermediate Staphylococcus aureus in a Pharmacokinetic/Pharmacodynamic Model

New antimicrobial agents and novel combination therapies are needed to treat serious infections caused by methicillin-resistant Staphylococcus aureus (MRSA) with reduced susceptibility to daptomycin and vancomycin. The purpose of this study was to evaluate the combination of ceftaroline plus daptomycin or vancomycin in an in vitro pharmacokinetic/pharmacodynamic model. Simulations of ceftaroline-fosamil at 600 mg per kg of body weight every 8 h (q8h) (maximum free-drug concentration in serum [fC_max], 15.2 mg/liter; half-life [t_1/2], 2.3 h), daptomycin at 10 mg/kg/day (fC_max, 11.3 mg/liter; t_1/2, 8 h), vancomycin at 2 g q12h (fC_max, 30 mg/liter; t_1/2, 6 h), ceftaroline plus daptomycin, and ceftaroline plus vancomycin were evaluated against a clinical, isogenic MRSA strain pair: D592 (daptomycin susceptible and heterogeneous vancomycin intermediate) and D712 (daptomycin nonsusceptible and vancomycin intermediate) in a one-compartment in vitro pharmacokinetic/pharmacodynamic model over 96 h. Therapeutic enhancement of combinations was defined as ≥2 log₁₀ CFU/ml reduction over the most active single agent. The effect of ceftaroline on the membrane charge, cell wall thickness, susceptibility to killing by the human cathelicidin LL37, and daptomycin binding were evaluated. Therapeutic enhancement was observed with daptomycin plus ceftaroline in both strains and vancomycin plus ceftaroline against D592. Ceftaroline exposure enhanced daptomycin-induced depolarization (81.7% versus 72.3%; P = 0.03) and killing by cathelicidin LL37 (P < 0.01) and reduced cell wall thickness (P < 0.001). Fluorescence-labeled daptomycin was bound over 7-fold more in ceftaroline-exposed cells. Whole-genome sequencing and mutation analysis of these strains indicated that change in daptomycin susceptibility is related to an fmtC (mprF) mutation. The combination of daptomycin plus ceftaroline appears to be potent, with rapid and sustained bactericidal activity against both daptomycin-susceptible and -nonsusceptible strains of MRSA.     

Vancomycin-loaded titanium coatings with an interconnected micro-patterned structure for prophylaxis of infections: an in vivo study

Titanium (Ti) and its alloys are widely applied as orthopedic implants for hip and knee prostheses, fixation, and dental implants. However, Ti and its alloys are bioinert and susceptible to bacteria and biofilm formation. Thus, surface biofunctionalisation of Ti is essential for improving the biofunction of Ti. The current in vitro study indicated that calcium phosphate bone cement with vancomycin doped on micro-patterned Ti with a grid-like structure surface could preserve the property of inhibition of bacterial adhesion and biofilm formation while not affecting the osteogenic differentiation. The present study investigated whether the biological performance of the bactericidal effect is preserved in vivo. The rabbit osteomyelitis model with tibial medullary cavity placement of Ti rods was employed to analyze the antibacterial effect of vancomycin-loaded Ti coatings with interconnected micro-patterned structure (TV). Thirty female rabbits (N = 10) were used to establish the implant-associated infection. Prior to implanting the T0 and TV rods into the medullary cavity of the left tibia of the rabbits, 10⁶ CFU mL⁻¹ methicillin-resistant Staphylococcus aureus (MRSA) was injected into the medullary cavity of the left tibia of the rabbits. The sterile Ti rod (NT) was used as the blank control. After 3 weeks, bone pathology was evaluated using X-ray and micro-CT. The in vivo study proposed that TV has the potential for prophylaxis against MRSA infection. Thus, the interconnected micro-patterned structured Ti rods loaded with vancomycin could be applied for preventing Ti implant-associated infections.

Vancomycin-loaded titanium coatings with an interconnected micro-patterned structure for prophylaxis of titanium implant associated infection.