Inducible nitric oxide synthase-deficient mice show exacerbated inflammatory process and high production of both Th1 and Th2 cytokines during paracoccidioidomycosis.

Paracoccidioidomycosis, the major systemic mycosis in Latin America, is caused by fungus Paracoccidioides brasiliensis. To analyze the influence of inducible nitric oxide synthase (iNOS) in this disease, iNOS-deficient (iNOS(-/-)) and wild-type (WT) mice were infected intravenously with P. brasiliensis 18 isolate. We found that, unlike WT mice, iNOS(-/-) mice did not control fungal proliferation, and began to succumb to infection by day 50 after inoculation of yeast cells. Typical inflammatory granulomas were found in WT mice, while, iNOS(-/-) mice presented incipient granulomas with intense inflammatory process and necrosis. Additionally, splenocytes from iNOS(-/-) mice did not produce nitric oxide, however, their proliferative response to Con-A was impaired, just like infected WT mice. Moreover, infected iNOS(-/-) mice presented a mixed pattern of immune response, releasing high levels of both Th1 (IL-12, IFN-gamma and TNF-alpha) and Th2 (IL-4 and IL-10) cytokines. These data suggest that the enzyme iNOS is a resistance factor during paracoccidioidomycosis by controlling fungal proliferation, by influencing cytokines production, and by appeasing the development of a high inflammatory response and consequently formation of necrosis. However, iNOS-derived nitric oxide seems not being the unique factor responsible for immunosuppression observed in infections caused by P. brasiliensis.

Deficiency of IL-12p40 subunit determines severe paracoccidioidomycosis in mice.

Paracoccidioidomycosis, the major systemic mycosis in Latin America, is caused by the thermally dimorphic fungus Paracoccidioides brasiliensis. To investigate the role of interleukin (IL)-12 in this disease, IL-12p40-/- deficient mice (IL-12p40-/-) and wild type mice (WT) were infected intravenously with viable yeast cells of P. brasiliensis 18 isolate. We found that, unlike WT mice, IL-12p40-/- mice did not control fungal proliferation and dissemination and succumbed to infection by day 21 after inoculation. Additionally, IL-12p40-/- mice presented a higher number of granulomas/mm2 in lung tissue than WT mice, and showed unorganized granulomas containing high numbers of yeast cells. Moreover, IL-12p40-/- mice did not release detectable levels of IFN-gamma, but they produced high levels of IL-10, as well as IgG1 antibody. Additionally, splenocytes from both infected IL-12p40-/- and WT mice exhibited a suppressed Con-A-induced T cell proliferative response. Our findings suggest that the IL-12p40 subunit mediates resistance in paracoccidioidomycosis by inducting IFN-gamma production and a Th1 immune response.

In vitro antifungal drug susceptibilities of dermatophytes microconidia and arthroconidia.

OBJECTIVES: Arthroconidia have been considered as the primary cause of infection by dermatophytes. However, the in vitro antifungal testing evaluates the responses mainly of microconidia or hyphae, and dermatophytes in vivo often produce arthroconidia, a cellular structure presumably more resistant to antifungals. The aim of this study was to compare the in vitro susceptibility of microconidia and arthroconidia of Trichophyton rubrum, Trichophyton tonsurans and Trichophyton equinum to griseofulvin, itraconazole, terbinafine, fluconazole, amphotericin B and hygromycin B. METHODS: Microconidia and arthroconidia were produced in vitro, and their susceptibility to each drug was evaluated by assessing the CLSI M38-A broth microdilution method. RESULTS: Arthroconidia of all strains analysed appeared to be more resistant to fluconazole, griseofulvin and itraconazole than microconidia. The MIC of terbinafine was the same for microconidia and arthroconidia for all strains, and the MIC of amphotericin B for microconidia and arthroconidia was the same for isolates of T. equinum and T. tonsurans, but differed for T. rubrum. Finally, the level of resistance of microconidia for all strains towards the antibiotic hygromycin B was from 25 to 400 mg/L. CONCLUSIONS: The difference in the susceptibility between microconidia and arthroconidia depends on the drug and on the strain, and may be one of the causes of therapeutic failure. Also, the level of resistance to the antibiotic hygromycin B presented by microconidia of these isolates will allow the use of hygromycin resistance as a dominant marker in fungal transformation procedures in future studies of gene function.