In Vitro Activities at pH 5.0 and pH 7.0 and In Vivo Efficacy of Flucytosine against Aspergillus fumigatus

Abstract

The antifungal agent flucytosine was found to be active in vitro against Aspergillus fumigatus isolates when the MIC was determined at pH 5.0 instead of pH 7.0. The in vitro MIC at pH 5.0 corresponded to the in vivo efficacy of flucytosine monotherapy in a murine model of invasive aspergillosis.The efficacy of flucytosine for the treatment of invasive aspergillosis (IA) has been controversial, although the evidence supporting its use for this condition is limited (5, 10, 11). Most guidelines do not recommend the use of the drug for the treatment of IA (1, 13), although use of the combination of amphotericin B and flucytosine for the treatment of Aspergillus osteomyelitis and joint infection was recommended previously (5).Most isolates of Aspergillus species are not susceptible to flucytosine, with MICs typically being Ͼ64 mg/liter (8). However, we previously demonstrated that the activity of flucytosine against Aspergillus species increased when the pH of the medium was lowered from 7.0 to 5.0 (8). Viviani et al. (12) showed that the in vitro activity of flucytosine against Cryptococcus neoformans at pH 5.4 correlated well with the clinical outcome. We investigated the flucytosine MIC distribution of Aspergillus fumigatus at pH 7.0 and pH 5.0 and determined which condition of MIC testing best corresponded with in vivo efficacy in a nonneutropenic murine model of IA.The in vitro activity of flucytosine against 50 clinical A. fumigatus isolates from our private fungus culture collection was determined by using the microdilution format of the CLSI (formerly the NCCLS) M38-A protocol (6). The isolates were cultured from clinical specimens from patients admitted to the Radboud University Medical Center and other Dutch hospitals between 1999 and 2005. All isolates were tested in duplicate at pH 7.0 and pH 5.0, and the MIC was read as the 50% inhibition of growth compared to the growth for the control. The pH was adjusted to 5.0 by using 100 mM citrate buffer.Female CD-1 outbred mice (weight, 20 to 29 g; Charles River Laboratories, Sulzfeld, Germany) were used. The animal studies were conducted in accordance with the recommendations of the European Community (Directive 86/609/EEC, 24 November 1986) and were approved by the institutional animal care and use committee of Radboud University. Inocula were prepared as conidial suspensions in saline containing 0.05% Tween 80, counted microscopically, and adjusted to the required concentration. Mice were infected with the 90% lethal dose (LD 90 ) by injection of 0.1 ml of the conidial suspension into the orbital vein.Flucytosine was dissolved in distilled water, according to the instructions of the manufacturer, and administered intraperitoneally. Treatment was begun 2 h after infection. Groups of 10 mice each were treated for 7 days with 100 mg/kg of body weight every 6 h (q6h), 100 mg/kg every 12 h (q12h), 200 mg/kg q6h, and 200 mg/kg q12h. Control mice were infected but received only distilled water. Animals were checked twice daily for clinical signs and mortality. Mort...

Efficacy of Posaconazole against Three Clinical Aspergillus fumigatus Isolates with Mutations in the cyp51A Gene

“…We conclude that our model supports the view that the efficacy of posaconazole treatment in A. fumigatus is dependent both on drug exposure and on the MIC for the isolate, as with other antifungal drugs (21,30). In addition, the quantitative relationship between exposure and effect is comparable to that found for Candida strains (1).…”

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Genetic Diversity and In Vitro Antifungal Susceptibility of 200 Clinical and Environmental Aspergillus flavus Isolates

“…Flucytosine was found to be active in vitro against Aspergillus isolates when the MIC was determined at pH 5.0 instead of pH 7.0 (15). In addition, the in vitro MIC at pH 5.0 corresponded to the in vivo efficacy of flucytosine monotherapy in a murine model of invasive aspergillosis (16,17).…”

Pharmacodynamics of Itraconazole against Aspergillus fumigatus in an In Vitro Model of the Human Alveolus: Perspectives on the Treatment of Triazole-Resistant Infection and Utility of Airway Administration

“…The reason for the discordance may be related to the use of an endpoint for MIC determination that does not require complete inhibition of growth. Furthermore, in vitro activity of flucytosine against Aspergillus is improved at a pH of 5 versus 7, which better predicts the activity that is observed with this compound in mice with disseminated aspergillosis (19,20). Using interaction modeling, the combination of aerosolized itraconazole and systemically administered 5FC is additive against an L98H mutant.…”