Efficacy of Iclaprim against Wild-Type and Thymidine Kinase-Deficient Methicillin-Resistant Staphylococcus aureus Isolates in an In Vitro Fibrin Clot Model

Abstract

Iclaprim is a novel diaminopyrimidine antibiotic that is active against methicillin-resistant Staphylococcus aureus (MRSA). However, it is known that the activity of diaminopyrimidines against S. aureus is antagonized by thymidine through uptake and conversion to thymidylate by thymidine kinase. Unlike with humans, for whom thymidine levels are low, thymidine levels in rodents are high, thus precluding the accurate evaluation of iclaprim efficacy in animal models. We have studied the bactericidal activity of iclaprim against an isogenic pair of MRSA isolates, the wild-type parent AW6 and its thymidine kinase-deficient mutant AH1252, in an in vitro fibrin clot model. Clots, which were aimed at mimicking vegetation structure, were made from human or rat plasma containing either the parent AW6 or the mutant AH1252, and they were exposed to homologous serum supplemented with iclaprim (3.5 g/ml), trimethoprim-sulfamethoxazole (TMP-SMX; 8/40 g/ml), vancomycin (40 g/ml), or saline, each of which was added one time for 48 h. In rat clots, iclaprim and TMP-SMX were bacteriostatic against the parent, AW6. In contrast, they were bactericidal (>3 log 10 CFU/clot killing of the original inoculum) against the mutant AH1252. Vancomycin was the most active drug against AW6 (P < 0.05), but it showed an activity similar those of iclaprim and TMP-SMX against AH1252. In human clots, iclaprim was bactericidal against both AW6 and AH1252 strains and was as effective as TMP-SMX and vancomycin (P > 0.05). Future studies of animals using simulated human kinetics of iclaprim and thymidine kinase-deficient MRSA, which eliminate the thymidine-induced confounding effect, are warranted to support the use of iclaprim in the treatment of severe MRSA infections in humans.

Synergistic interaction between phage therapy and antibiotics clears Pseudomonas aeruginosa infection in endocarditis and reduces virulence

“…Experiments were done in triplicates. Plasma clots were prepared in 96-well microplates, as previously described [ 16 ]. Phage titers were measured by plaque assays [ 15 ].…”

Antimicrobial treatment of nosocomial meticillin-resistant Staphylococcus aureus (MRSA) pneumonia: current and future options

“…Additionally, resistance is less likely to develop as a result of the use of other MRSA antimicrobials, as iclaprim has a different mechanism of action than many of the agents that are currently available or in development. However, the antibacterial effect of DHFR inhibitors can be antagonised in vitro by supplementing thymidine owing to bacterial uptake of exogenous thymidine and its subsequent conversion into thymidine monophosphate by thymidine kinase, bypassing the DHFR inhibition [121,122]. Because thymidine is found in large concentrations in human pus, some believe that iclaprim and other DHFR inhibitors are less effective in purulent infections [123].…”

Defeating Antibiotic- and Phage-Resistant Enterococcus faecalis Using a Phage Cocktail in Vitro and in a Clot Model

“…A recognized tool for the study of bacterial endocarditis ( Hershberger et al, 2000 ), we used the in vitro fibrin-clot infection model ( McGrath et al, 1994 ; Entenza et al, 2009 ) to assess the efficacy of the cocktail and the individual phages EFDG1 and EFLK1 against E. faecalis V583 and EFDG1 r ( Figure 5 ). CFU/ml results following a 6-h incubation of E. faecalis V583 with EFLK1 phage and cocktail#1 showed a CFU reduction of three logs with EFDG1 phage and six logs with EFLK1 phage and cocktail#1.…”

“…The in vitro fibrin clot model was prepared according to the protocol described by McGrath et al (1994) and Entenza et al (2009) . Overnight cultures (10 9 CFU/ml) of E. faecalis individually or in a mixed cultures of naïve and resistant bacteria were diluted 1:10 with citrated plasma.…”