Dynamic changes in the morphology of Cryptococcus neoformans during murinepulmonary infection

The pathogenesis of Cryptococcus neoformans infection has been studied exte nsively with respect to inflammatory and pathological changes, but very lit tle information is available regarding the morphology of yeast cells during the course of infection. Electron microscopy of Cryptococcus neoformans in murine pulmonary infection revealed increased cell wall thickness with tim e, but this difference was only partially accounted for by increases in cel l diameter. Cell walls of melanized cells were thicker than those of nonmel anized cells 2 h after infection, and the cell wall of yeast became blacker with time, suggesting that melanization contributes to the increased cell wall thickness. Heterogeneous cell populations emerged, with the appearance of giant forms. While for C. neoformans ATCC strain 24067 (serotype D) the full spectrum of cell sizes were observed, for strains H99 (serotype A) an d 3501 (serotype D) cells were divisible into two populations, giant and mi cro forms. In contrast to cellular heterogeneity, the epitope recognized by a protective mAb on the capsular glucuronoxylomannan (GXM) was found at al l times of infection. Immunoelectron microscopy using mAbs to GXM demonstra ted reactivity with intracellular structures, suggesting that synthesis of capsular polysaccharide occurs, at least in part, in the cytoplasm. In summ ary, the results indicate that: (i) the infection is dynamic with respect t o yeast cell morphology; (ii) giant cell forms arise in tissue during the c ourse of infection; (iiii) cell walls blacken and thicken during the course of infection, consistent with melanin synthesis during infection; and (iv) GXM epitopes are found in the capsule, cell wall and cytoplasm, consistent with intracellular polysaccharide synthesis. The results indicate that the population of C. neoformans cells in tissue is in a highly dynamic state, implying that the immune system must confront cells with varying characteri stics during the course of infection.