Penicillium marneffei is a dimorphic pathogenic fungus that can cause disseminated and progressive disease. The disease is one of the most common infections in AIDS patients in endemic area, which is Southeast Asia and China, especially in northern Thailand

in the latter it is the third common infection after tuberculosis and cryptococcosis. The exact route of infection in humans is not known, but it is assumed that infection is acquired by inhalation of the conidia. Previous studies using gel electrophoresis and immunoblot assays demonstrated the presence of humoral immune responses to this fungus in sera from 33 AIDS patients with culture-confirmed penicilliosis marneffei. Four major immunogenic proteins of 200, 88, 54 and 50 kDa were produced in large quantities from P. marneffei yeast-form during the deceleration and early stationary phases of growth. Immunoreactivity of these proteins was detected in 72.7, 93.9, 60.6 and 57.6% of patients, respectively. The bands of 88, 54 and 50 kDa gave strong reactions with half of serum samples. The present study has confirmed this work by using the same method. The culture filtrate protein antigens were prepared from the yeast phase of P. marneffei 391H and 302BM. Their antigenicity was tested by immunoblot assay with pooled sera of 10 AIDS patients infected with P. marneffei. It was found that the 88 and 50 kDa proteins were highly immunogenic and secreted in large amounts into the culture filtrate of P. marneffei (yeast-form). Both antigens were isolated and purified by using preparative polyacrylamide gel electrophoresis. The 88 kDa protein was isolated as a single band in fractions 160-210 by using Prep-cell preparative gel electrophoresis. However, after these fractions were pooled and concentrated by using ultrafiltration and a centricon device, the sample was contaminated with some lower molecular mass proteins. The 50 kDa protein was isolated as a single band by using Mini-Protean II preparative gel electrophoresis following with Coomassie brilliant blue staining, band excision and electro-elution. Two-dimensional gel electrophoresis was used to determine whether these antigens were single or multiple proteins. The proteins were separated according to isoelectric points in the first dimension and according to molecular weight in the second dimension. The results revealed that the 88 and 50 kDa proteins had isoelectric point values of approximately (+,ฃ) 4.5 to 5.6 and (+,ฃ) 4.5 to 5.1, respectively

the data did not indicate whether they were single or multiple proteins. These separated proteins were then transferred onto nitrocellulose membrane to study their antigenicity and concanavalin A and wheat germ agglutinin binding properties. The results indicated that both antigens were glycoproteins. The 88 kDa antigen has higher affinity for concanavalin A than the 50 kDa antigen. They were characterized as mannoproteins. Both antigens did not bind wheat germ agglutinin.