Avaliação da resposta humoral a antígenos de Lacazia loboi utilizando sorosde pacientes com lacaziose

Abstract

Jorge Lobos disease is a mycosis of the skin and subcutaneous tissue caused by Lacazia loboi, a fungus that presents phenotypic similarities to Paracoccidioides brasiliensis. Because it resists culture, research to characterize and isolate its DNA and antigenic proteins has been a problem. Thence, most previous serological studies have used antigens from P. brasiliensis. The objective of the present study isto evaluate the host humoral immune response to L. loboi yeast-like cells extracted from mice experimentally infected with the fungus. Herein, L. loboi antigens were obtained from an experimentally infected animal. BALB/c mice were inoculated with yeast-like cells extracts obtained from fragments of skin lesions of patients with lacaziosis. Six months after inoculation, the mice developed typical lesions in both foot pads. Mice were sacrificed and the lesions from the foot pads were excised, thenmacerated in a glass tissue grinder and the fungal suspension was filtered to eliminate debris. The extracted antigens were maintained at 20ºC. P. brasiliensis gp43 glycoprotein was isolated from the Pb18 strain and purified from the concentrated crude exoantigen. The final solution was loaded onto a column of CNBr-4B coupled to monoclonal antibody gp43 17c for affinity chromatography. Western blotting analyses were carried out using sera from 24 patients with lacaziosis, five patients with paracoccidioidomycosis and sera from five healthy controls from a non endemic area. The same procedure was performed with serafrom six infected and nine non infected mice. IgG antibodies from all patients and mice with lacaziosis detected a ~193kDa antigen. The purified gp43 glycoprotein of P. brasiliensis was detected by the IgG of all evaluated sera, but the IgG in these sera failed to detect the same molecular antigen in the extracts from L. loboi yeastlike cells. Sera from healthy volunteers and control mice did not react with the antigens used. The molecular characterization of the detected antigens, particularlythe ~193kDa protein, may be important for the development of new treatments, immunotherapy, vaccine and diagnostic tests for lacaziosis.