Experimental paracoccidioidomycosis in hamsters (Mesocricetus auratus): Gestational interactions

Paracoccidioides brasiliensis is a dimorphic fungus presenting specific steroid hormone receptors, both in the yeast and mycelial forms and estrogen inhibits the transition from mycelium to yeast. In the acute phase, the disease occurs with equal frequency in both sexes but in adults, females are spared. Placental fungal infection has been reported, but references to fetal infection have not been confirmed. We used 78 Syrian female hamsters divided into 3 groups: GI consisted of 30 infected mated females, GII of 20 infected unmated females and GIII of 28 uninfected mated females. Animals of group I were mated 4 weeks after infection and half of them were submitted to cesarean section on day 15 after successful mating; the other half was maintained and submitted to cesarean section and sacrificed 14 weeks after infection. Half of the animals of group II were sacrificed seven weeks and the other half 14 weeks after infection. Uninfected animals of group III were treated the same as the animals of group I. The animals were infected with strain 18 of P. brasiliensis by the intracardiac route. We evaluated the disease by the volume of granulomas in different organs, number of fungi in liver and spleen and the immunologic responses [ELISA, Double Immunodifusion (DID), Delayed Hypersensitivity Skin Test (DHT) and Macrophage Migration Inhibition (MMI)]. We studied the infection through the gestation by evaluation of the abortions, morphologic and clinic examinations of the fetuses. Our results showed that the infection did not transfer to the fetus through the placenta, but the number of abortions was larger among infected females. The newborns of GI females were smaller, weighed less and showed little vitality. The disease was more severe and disseminated in infected mated females, especially in the second sacrifice 14 weeks after inoculation, when the total volume of granulomas in them (56.3 mm) was much greater than in the infected unmated females (12 mm).

Effect of cytokines on the in vitro fungicidal activity of monocytes from paracoccidioidomycosis patients

Peripheral blood monocytes obtained from paracoccidioidomycosis patients and healthy individuals were preactivated with recombinant gamma interferon (IFN-gamma) in different concentrations (250, 500 and 1000 U/ml) and evaluated for fungicidal activity against Paracoccidiodes brasiliensis strain 18 (Pb 18, high-virulence strain) and strain 265 (Pb 265, low-virulence strain) by plating of cocultures and counting of colony-forming units, after 10 d. Monocytes from healthy individuals failed to present fungicidal activity against P. brasiliensis even after IFN-gamma activation at the three concentrations. However, patient, monocytes activated with IFN-gamma (1 000 U/ml) showed a significant fungicidal activity when compared to that obtained with non-activated or activated cells with other IFN-gamma concentrations (250 and 500 U/ml). Moreover,,patient monocytes presented higher fungicidal activity than the control, even before the activation process. These results may be explained by the activation state of patients' cells as a function of the in vivo contact with the fungus, which was confirmed by their higher capacity to release H2O2 in vitro. Unlike the results obtained with Ph 18, patient and control cells presented a significant fungicidal activity against Pb 265, after priming with IFN-gamma. These results are explained by the higher levels of TNF-alpha in supernatants of cultures challenged with Pb 265. Moreover, higher levels of the cytokine were obtained in patient cell supernatants. Taken together, our results suggest that for effective killing of P. brasiliensis by monocytes, an initial activation signal induced by IFN-gamma is necessary to stimulate the cells to produce TNF-alpha. This cytokine may be involved, through an autocrine pathway, in the final phase activation process. The effectiveness of this process seems to depend on the virulence of the fungal strain and the activation state of the challenged cells. (C) 2003 Editions scientifiques et medicales Elsevier SAS. All fights reserved.

POLYSEROSITIS IN A PATIENT WITH ACUTE PARACOCCIDIOIDOMYCOSIS AND HEPATOSPLENIC SCHISTOSOMIASIS

A severe case of juvenile paracoccidioidomycosis (PCM), manifested as cholestatic jaundice, lymphnode enlargement and an unusual form of polyserositis, associated with portal hypertension secondary to schistosomiasis, as well as bacteremias caused by E. coli and S. aureus and post-transfusional hepatitis C is reported. Temporary unresponsiveness of in vivo and in vitro cellular immune responses to P. brasiliensis were registered. The authors discuss the possible interference of either agent in the host immune response, thus explaining the severity of PCM in the present case.

DELAYED-TYPE HYPERSENSITIVITY RESPONSE IN AN ISOGENIC MURINE MODEL OF PARACOCCIDIOIDOMYCOSIS

The specific delayed-type hypersensitivity (DTH) response was evaluated in resistant (A/SN) and susceptible (B10.A) mice intraperitoneally infected with yeasts from a virulent (Pb18) or from a non-virulent (Pb265) Paracoccidioides brasiliensis isolates. Both strains of mice were footpad challenged with homologous antigens. Pb18 infected A/SN mice developed an evident and persistent DTH response late in the course of the disease (90th day on) whereas B10.A animals mounted a discrete and ephemeral DTH response at the 14th day post-infection. A/SN mice infected with Pb265 developed cellular immune responses whereas B10.A mice were almost always anergic. Histological analysis of the footpads of infected mice at 48 hours after challenge showed a mixed infiltrate consisting of predominantly mononuclear cells. Previous infection of resistant and susceptible mice with Pb18 did not alter their DTH responses against heterologous unrelated antigens (sheep red blood cells and dinitrofluorobenzene) indicating that the observed cellular anergy was antigen-specific. When fungal related antigens (candidin and histoplasmin) were tested in resistant mice, absence of cross-reactivity was noted. Thus, specific DTH responses against P. brasiliensis depend on both the host's genetically determined resistance and the virulence of the fungal isolate.

The use of glucan as immunostimulant in the treatment of paracoccidioidomycosis

A group of 10 patients, nine of them seriously infected with Paracoccidioides brasiliensis (G1), received glucan (beta-1,3 polyglucose) as an immunostimulant intravenously once a week for one month, followed by monthly doses (10 mg) over an ii-month period, together with a specific anti-fungal agent as an immunostimulant. A second group of eight moderately infected patients (G2) was treated with only the anti-fungal agent. Among the patients in G1, there was only one case of relapse compared with five in G2. Values for the erythrocyte sedimentation rate (ESR) showed a significant difference (P < 0.01) post-treatment in G1 patients, when compared with the pretreatment levels. There was also a significant reduction (P < 0.001) in the level of serum antibodies to P. brasiliensis in the G1 patients in post-treatment examinations. The phytohemagglutinin (PHA) skin test showed a positive reaction among the patients in G1 (P < 0.01) post-treatment and there was a tendency towards an increase in the number of CD4+ T lymphocytes in both groups after treatment. The serum level of tumor necrosis factor (TNF) proved to be significantly higher (P < 0.02) in the G1 patients during treatment. In the G1 patients, the correlation between ESR and TNF tended to be negative whereas that between ESR and serum antibodies was positive. The present results indicate that the patients who received glucan, in spite of being more seriously ill, had a stronger and more favorable response to therapy.

Oral paracoccidioidomycosis: a study of 36 South American patients

Paracoccidioidomycosis (South American blastomycosis) is an uncommon, progressive systemic mycosis, virtually only seen in persons who have visited Latin America. Reports of oral lesions are extremely rare in the English-language literature. Thirty-six adults with oral lesions as the first sign of paracoccidioidomycosis are described; this appears to be the largest series in the dental literature. All had chronic proliferative mulberry-like ulcerated oral lesions; the diagnosis was confirmed histologically. The gingiva or alveolar process was the typical site, but lesions were also seen particularly on the palate and lip. Most of the patients proved to have detectable pulmonary involvement. Patients with lesions in the oropharynx, tongue, or floor of mouth all had confirmed pulmonary lesions.

Imbalance of IL-2, IFN-gamma and IL-10 secretion in the immunosuppression associated with human paracoccidioidomycosis

Patients with paracoccidioidomycosis (PCM) display a certain degree of immunecompromise characterized by lymphocyte hyporesponsiveness to the main Paracoccidioides brasiliensis antigen (gp43). To determine whether cytokines are involved in this state, we evaluated the secretion of IL-2, IL-10 and IFN-gamma by peripheral blood mononuclear cells (PBMC) from patients with the acute (AF) and chronic (CF) forms of PCM and from healthy, P. brasiliensis-sensitized subjects. gp43-stimulated PBMC from healthy subjects produced substantial amounts of IL-2, IFN-gamma and IL-10, whereas PBMC from AF and CF patients produced low levels of IL-2 and IFN-gamma but substantial amounts of IL-10, Phytohaemagglutinin-induced cytokine secretion was comparable among AF and CF patients and healthy subjects, suggesting integrity of non-specific cellular immune mechanisms in PCM. gp43-pulsed adherent cells, but not non-adherent cells, mere the main source of IL-10, Moreover, IL-2 and IFN-gamma secretion correlated inversely with the amount of specific antibodies produced by patients and healthy subjects. Our results suggest that the imbalance in cytokine production of patients with PCM plays a role in the gp43-hyporesponsiveness and the marked (non-protective) antibody production of these patients. (C) 2001 Academic Press.