A total of 60 clinical isolates of Paracoccidioides brasiliensis were tested for susceptibility to sulfadiazine and sulfadimethoxyne by the agar dilution technique. A modification of the Mueller-Hinton medium was devised which gave good growth of the yeast form. The minimum inhibitory concentrations for only 51.6% of the isolates were in the range of the recommended blood serum concentration (50 ,ug/ml (7,14).In spite of these observations, there has been no survey of the in vitro susceptibility of the causative agent, Paracoccidioides brasiliensis, to sulfonamides. Earlier studies either employed complex culture media, which could have antagonized the effect of the drug, or made use of the fungus mycelial phase, which is different from the phase found in tissues, that is, the yeast phase of this dimorphic fungus (5,8,15 This report describes the procedure developed for sulfonamide susceptibility testing and presents data on the susceptibility pattern of 60 different isolates of P. brasiliensis to sulfadiazine and sulfadimethoxyne.
MATERIALS AND METHODSFungi. Sixty-five isolates of P. brasiliensis were used in this study. All were from clinical sources (Colombian patients) and represented the original isolate at time of diagnosis, with the exception of three which were obtained during relapses. The stock cultures were kept in Sabouraud glucose agar at approximately 25°C. When needed, they were converted to the yeast phase by subculturing in the special media described below and incubated at 36°C. Identity confirmation was obtained by demonstration of dimorphism and by the characteristic microscopical appearance, multiple budding yeast cells, in cultures incubated at 360C. Culture media. It was necessary to find a medium which would not antagonize sulfonamides and which, at the same time, could promote abundant growth of P. brasiliensis. Earlier attempts to culture the fungus in the conventional Mueller-Hinton medium (BBL Microbiology Systems, Cockeysville, Md.) used for sulfonamide susceptibility testing of bacteria, proved unsuccessful (4, 22). A series of experiments was performed by enriching such a medium with various chemicals known to promote growth in the fungus (1,17