In vitro susceptibility testing of Paracoccidioides brasiliensis to sulfonamides

Abstract

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

Anti-fungal Activity of Sulfamethoxazole toward Aspergillus Species

“…16) Moreover, it has been reported that the combination of SMX and TMP acts synergistically against the dimorphic fungus Paracoccidioides brasiliensis. 17,18) Those reports suggest that SMX-TMP acts toward fungus. In the present study ( Fig.…”

Paracoccidioidomycosis in children: clinical presentation, follow-up and outcome

“…Moreover, it is inexpensive, may be infused intravenously in the severe forms of PCM, it is easy to administer orally and it is available in public health services. According to some, failures of treatment with this association are mostly secondary to poor patient compliance 19,24,27,30 .…”

Influence of Different Media, Incubation Times, and Temperatures for Determining the MICs of Seven Antifungal Agents against Paracoccidioides brasiliensis by Microdilution

“…MMH medium is not a chemically defined medium, and microorganisms can obtain folic acid or its precursors from different sources. MMH medium was tested in our study because it was previously used in MIC assays with agar dilution for P. brasiliensis isolates (23), yielding values similar to those obtained in RPMI 1640 medium, and because yeast morphology was maintained at 37°C in this medium until the end of the protocol. Our results showed that the MICs with MMH medium were similar to those found with RPMI 1640 medium when sulfamethoxazole and the sulfamethoxazole-trimethoprim combination were tested.…”

“…The standard RPMI 1640 medium at 34.54 g per liter was buffered with 0.165 M morpholinepropanesulfonic acid (MOPS) at pH 7.0. MVM medium was prepared as recommended by Restrepo and Jimé-nez (30), and MMH medium (Mueller-Hinton medium supplemented with 10 g/liter of glucose, 5 g/liter of ammonium sulfate, and 10 mg/liter of thiamine [Sigma-Aldrich]) was prepared as recommended by Restrepo and Arango (23). All drugs were tested in these three media for all of the isolates.…”

“…These studies have generally used modified versions of the NCCLS M27-A2 protocols or a Shadomy modified protocol (10). These modifications include differences in inoculum preparation, media used in MIC assays, incubation periods, and experimental temperatures (5,6,(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23). However, standardization of the methods used for in vitro susceptibility testing is important to facilitate the establishment of interpretative breakpoints and quality control parameters.…”