The oral route in the pathogenesis of paracoccidioidomycosis: an experimental study in BALB/c mice infected with P. brasiliensis conidia.

Due to the high frequency of oral mucosal lesions observed in paracoccidioidomycosis patients, it was advocated that the infection was acquired by the traumatic implantation of the etiologic agent Paracoccidioides brasiliensis. Although at present this theory is considered invalid, it has not yet been excluded in experimental studies. In order to determine if intra-oral inoculation could explain the pathogenesis of paracoccidioidomycosis, 64 BALB/c mice were inoculated intra-orally with 850.000 viable P. brasiliensis conidia into the mandibular body. Animals were sacrificed at various time intervals up to 20 weeks and cultures were made from gingiva, lungs, spleen, and liver. Additionally, histopathological studies of the mandibular body were also performed. P. brasiliensis was isolated from all gingival tissues during the interval 24-72 h, indicating that the infection was active. During the 5-10 week period, the infection appeared to have been controlled at the inoculation site as cultures showed a significant reduction in colony forming units (CFU); however, at the 15-20 week period such control was lost and the fungus was recovered once more. Dissemination to other body sites was rare; thus, the lungs were involved in just one animal (2%), the liver in two (3%) and the spleen in seven (11%). The infection became established as proven by positive organ cultures, but the dissemination pattern did not correspond to the one observed in humans. Based on these findings, the intra-oral traumatic route does not appear to mimic the natural history of paracoccidioidomycosis.

Activity of two different triazoles in a murine model of paracoccidioidomycosis.

A new orally absorbable triazole (Schering 39304) with a long serum half-life in man (60 hours), was tried in a murine model of progressive paracoccidioidomycosis and compared with itraconazole, another triazole which has proven effective in this mycosis. Only 15% of the infected, untreated mice survived while 53 to 75% of the animals receiving itraconazole survived. Mice treated with Schering 39304 exhibited higher (86-100%) survival rates. Statistically, the 5 mg/kg Sch 39304 was superior to the 50 mg/kg itraconazole dose. Lung cultures showed that 20 mg/kg/day of Sch achieved sterilization of the infectious foci. These results indicate that the new triazole will have a place in the treatment of paracoccidioidomycosis.

Water as a substrate for the development of Paracoccidiodes brasiliensis mycelial form.

Two isolates of P. brasiliensis in the mycelial form were studied for their capacity to survive and grow in sterile distilled water (SDW). Inoculum for the experiments consisted of a spectrophotometrically-standardized suspension of washed and homogenized mycelial fragments; these had been obtained from 2-week old cultures grown in a synthetic medium (SM). Series of tubes with SDW and SM were incubated with the above suspension and kept stationary for 6 months at either 4 degrees C or room temperature (RT). Growth was measured by dry weight (DW) and turbidity (OD) determinations; additionally, CFU and ultrastructural appearance by transmission electron microscope (TEM) were assessed for one of the isolates. In general, cultures in SM at RT, grew exponentially after 2 weeks, becoming stationary for 7 weeks and then, declining abruptly. In SDW, fungal development was slow for 5 months when an increase in mass was recorded. When incubated at 4 degrees C, both SDW and SM cultures required longer time to develop but mass also increased. Morphologically, mycelial elements in SDW at RT exhibited increased lipid vacuoles and glycogen deposits but were otherwise normal up to 6 weeks when they presented the inter-hyphae-hyphae phenomenum. In SDW P. brasiliensis appears to utilize debris from its degenerated fungal partners to continue growing.

Pathogenesis of paracoccidioidomycosis: a histopathological study of the experimental murine infection.

The pathogenesis of primary pulmonary P. brasiliensis infection, the systemic dissemination which followed, and the histopathology of the main organs involved was studied in a murine model of chronic paracoccidioidomycosis. Adult male BALB/C mice, were challenged intranasally with 26 X 10(-6) viable P. brasiliensis yeast cells. We inoculated 86 animals which were sacrificed from 0 h to 20 weeks. As controls, 11 mice were instilled with saline solution, and 48 with 26 X 10(-6) heat-killed. P. brasiliensis yeast cells. None of the animals receiving saline, exhibited pathologic alterations; 11.6% of those inoculated with the heat-killed cells, revealed mild, transitory acino-pulmonary neutrophilic infiltrates. The animals infected with viable cells, developed a systemic process affecting mainly the lungs (46.5%), liver (18.6%), lymph-nodes (18.6%), and spleen (3.5%). In this group of animals, lung lesions were detected regularly at all time periods from 3 h to 20 weeks. A multiple bronchopneumonic process was initially observed at 6 h, reached its maximum intensity around the third day, subsided thereafter but did not disappear and reactivated after the fifth week to become stationary until the end of experiments. Dissemination to other organs occurred early, and apparently by the hematogenous route. Initially the inflammatory cell infiltrate was mainly neutrophilic. With time, these cells were gradually replaced by lymphocytes, histiocytes and plasmocytes. Granuloma configuration of the cell infiltrate was distinctly seen around the fifth week, with multinucleated giant cells appearing at the ninth week. Hiliary lymph-node involvement was rare (7%) and primary lung lesions, as seen in tuberculosis and histoplasmosis, were not observed.

Itraconazole therapy in lymphangitic and cutaneous sporotrichosis.

Itraconazole, a new orally absorbable azole derivative, was used for the treatment of 17 patients with cutaneous and lymphangitic sporotrichosis. The drug, administered at a dose of 100 mg/day, proved to be effective in all cases. Lesions disappeared and cultures became negative after 90 to 180 days of therapy. There were no major side effects. Posttherapy evaluations, done in 14 of 17 cases for an average of 115 days, revealed no relapses. Objective evaluation of the treatment by means of a scoring system indicated complete resolution of the pretherapy abnormalities at varying periods; thus, 35.3% (six of 17) of the patients had recovered by 90 days, 45.4% (five of 11) by 120 days, and 83.3% (five of six) by 150 days of therapy. Consequently, therapy with itraconazole is an adequate alternative to iodide treatment in sporotrichosis.

Characteristics of the conidia produced by the mycelial form of Paracoccidioides brasiliensis.

The sporulation capacities of the mycelial form of Paracoccidioides brasiliensis were determined. Five different culture media were used and four human isolates studied. Conidia were produced in three agar media, namely water-agar, glucose-salts and yeast-extract. Corn meal and Sabouraud dextrose agars failed to induce sporulation. Various types of spores were characterized with peculiar bulging arthroconidia and single-celled, pear-shaped conidia predominating. The size of these conidia varied from 3.6 to 4.6 micron in length. It is concluded that the mycelial form of P. brasiliensis produces characteristic spores if the proper culture media are employed.

A comparison of mycelial filtrate - and yeast lysate - paracoccidioidin in patients with paracoccidioidomycosis.

A cytoplasmic antigen was obtained from P. brasiliensis yeast cells by water lysis and sonication, with the aim of detecting delayed hypersensitivity to the fungus. This antigen was studied in patients with paracoccidioidomycosis (active and inactive), tuberculosis and histoplasmosis as well as in normal individuals, and it was compared with a mycelial filtrate antigen employed in the past for the same purpose. The yeast paracoccidioidin proved superior to the mycelial preparation (62.0 vs 24.1%) in patients with active paracoccidioidomycosis; however, in inactive cases, both preparations gave similar results. The size of the reactions was also comparable for both products. In normal subjects and in patients with tuberculosis, skin reactivity was low (not over 10%) and within the expected range of the area. In contrast, patients with histoplasmosis proved highly reactive to both antigens. These results indicate that the newer paracoccidioidin has advantages over the mycelial product as it detects a higher proportion of reactors and as such, may be helpful in studies aimed at defining areas of endemicity in countries where paracoccidioidomycosis is present.