Propagation of Pneumocystis carinii in Vitro

Abstract

Summary: Pneumocystis carinii was propagated in vitro with primary embryonic chick epithelial lung (CEL) cells. Viability and growth of the organism were demonstrated by direct observation of the reproductive cycle in the Sykes-Moore chamber, serial passage with an increase in the number of mature cyst forms, the cytopathic effect of the organism on cell culture, and inhibition of growth of the organism by specific antiserum and pentamidine isethionate. Attempts to culture P. carinii indefinitely were not successful. However, cyst forms derived from murine and human sources increased 100-fold and 10-fold, respectively, during CEL cell culture passages. Serial passage of trophozoites alone resulted in the development of typical CPE and a maximum number of 2.8 10³ cyst forms. Autoradiographic methods demonstrated active DNA, RNA, and protein synthesis within the cyst and suggest that metabolic interaction between the host cells and the organisms occurred. The nature of the attachment of P. carinii to the host CEL cell was clearly discernible by scanning electron microscopy (SEM). In the reproductive cycle a vegetative cell (designated “trophozoite”) attached by tubular expansions to the host CEL cell, probably for the transport of essential nutrients, and then detached without entering the cell. Sporozoites developed within the detached young cyst, reaching a maximum number of eight within the mature cyst. Excystment occurred through single or multiple sites in the cyst wall, after which the released trophozoite attached to a new host cell. Speculation: The in vitro propagation of P. carinii provides a basis for further studies to characterize the biologic and biochemical features of this organism and a source of antigen for the development of serologie tests for specific antibody by which the epidemiology and spectrum of clinical disease can be delineated more clearly.