Radiometric detection of metabolic activity of Paracoccidioides brasiliensis and its susceptibility to amphotericin B and diethylstilbestrol

Paracoccidioidomycosis (South American blastomycosis) is a systemic disease, strikingly more frequent in males, caused by the dimorphic fungus Paracoccidioides brasiliensis. A radiometric assay system has been applied to study the metabolic activity and the effect of drugs on this fungus "in vitro". The Y form of the yeast, grown in liquid Sabouraud medium was inoculated into sterile reaction vials containing the 6B aerobic medium along with 2.0 μCi of 14C-substrates. Control vials, prepared in the same way, contained autoclaved fungi. To study the effects of amphotericin B (AB) (0.1 and 10 μg/ml) and diethylstilbestrol (DSB) (1.0, 5.0 and 10 μg/ml) extra controls with live fungi and no drug were used. All vials were incubated at 35°C and metabolism measured daily with a Bactec instrument. 14CO2 production by P. brasiliensis was slow and could be followed for as long as 50 days. AB at 10mg/ml and DSB at 5 μg/ml inhibited the metabolism and had a cidal effect on this fungus. The results with DSB might explain the low incidence of the disease in females. This technique shows promise for studying metabolic pathways, investi gating more convenient 14C-substrates to expedite radiometric detection and for monitoring the effects of other drugs and factors on the metabolism of P. brasiliensis "in vitro".

Paracoccidioidomycosis: a sequential histopathologic study of lesions in experimentally-infected rats

Female albino rats were used for the sequential histopathological study of experimental paracoccidioidomycosis. The animals were inoculated intraperitoneally with a strain of Paracoccidioides brasiliensis in the yeast-like phase, and sacrificed at given intervals from 1 to 168 days after inoculation; each animal received an inoculum of 4 x 10(6) cells in 0.8 ml of saline. The control group received saline containing scrapings of the culture medium. Tissue from the inoculation site was examined. The cellular population, the extracellular matrix, and the presence and characteristics of fungi were analysed in the inflammatory granulomatous process by light microscopy. The results allowed to separate the kinetic of the inflammatory response into three stages: 1) neutrophilic or macrophagic-neutrophilic; 2) pre-granulomatous; 3) granulomatous. Synthesis of the extracellular matrix began with the depositing of fibrin-like material, and increased gradually with deposits of collagen, proteoglycans, and glycoproteins. Parasites were present in all of the examined periods. Recurrences of the disease were clearly shown through the concurrence of recently-formed granulomas with older granulomas, implying that this type of granulomatous process does not eliminate the disease, nor is it able to limit fungal dissemination over a prolonged period of time.