Cryptococcus neoformans: intracellular or extracellular?

Response from Feldmesser et al.We appreciate the comments of Dr Levitz on our article; given his many contributions to our understanding of the interaction of Cryptococcus neoformans with human cells, his comments carry great authority. Although we are in general agreement with his letter, we would like to comment on some of the points he has raised.Dr Levitz suggests that the pathogenesis of human and murine infection might differ. We agree that caution is important when extrapolating from mice to humans. However, the histopathology of cryptococcosis in mice is very similar to that found in humans, and there is evidence for hueco cell formation in human tissue 1xPrimary cutaneous cryptococcosis: review and morphologic study. Noble, R.C. and Fajardo, L.F. Am. J. Clin. Pathol. 1971; 57: 13–22See all References1. Additionally, we are not sure that mice are more susceptible to cryptococcosis than humans, as experimental infection models in mice routinely use inocula that are several orders of magnitude greater than would be expected in naturally occurring human infections.The second paradox that Dr Levitz addresses is the preponderance of extracellular organisms in tissue sections from immunocompromised patients. A hallmark of cryptococcosis is the occurrence of large masses of organisms in tissue, often with little or no inflammation. Similar findings are seen in murine tissue with progressive infection. This paradox can be rephrased as a question: how can C. neoformans be a facultative intracellular pathogen when the tissue is full of extracellular organisms? The answer lies in the word ‘facultative’. C. neoformans can grow both intracellularly and extracellularly, but budding-index determinations indicate that the rate of intracellular growth is fivefold greater than extracellular growth 2xCryptococcus neoformans is a facultative intracellular pathogen in murine pulmonary infection. Feldmesser, M. et al. Infect. Immun. 2000; 68: 4225–4237Crossref | PubMed | Scopus (222)See all References2. This makes sense, as the nutritional environment for a yeast cell inside a dying macrophage is likely to be significantly richer than that experienced by a yeast cell in a mass of organisms. Hence, we agree that C. neoformans is both an intracellular and extracellular pathogen, but propose that many of the extracellular organisms arose from intracellular growth followed by liquefaction and digestion of host cells.The C. neoformans capsule is generally considered to be anti-phagocytic because phagocytic cells do not ingest yeast cells in the absence of complement or antibody opsonins in vitro. However, encapsulated C. neoformans cells are rapidly ingested in vivo, presumably owing to the presence of opsonins in tissue and/or differences in receptor expression for phagocytic cells in vivo. Studies in mice and rats, together with limited pathological studies of tissue from immunocompetent humans, indicate that C. neoformans is frequently found inside macrophages in tissue despite the presence of polysaccharide capsules in yeast cells 2xCryptococcus neoformans is a facultative intracellular pathogen in murine pulmonary infection. Feldmesser, M. et al. Infect. Immun. 2000; 68: 4225–4237Crossref | PubMed | Scopus (222)See all References, 3xPersistent Cryptococcus neoformans pulmonary infection in the rat is associated with intracellular parasitism, decreased inducible nitric oxide synthase expression, and altered antibody responsiveness to cryptococcal polysaccharide. Goldman, D.L. et al. Infect. Immun. 2000; 68: 832–838Crossref | PubMed | Scopus (81)See all References, 4xPrimary pulmonary cryptococcosis. Bahr, R.D. et al. Am. J. Roentgenol. Radium Ther. Nucl. Med. 1962; 87: 859–864PubMedSee all References, 5xTissue changes and tissue diagnosis in cryptococcosis. Baker, R.D. and Haugen, R.K. Am. J. Med. 1955; 25: 14–24See all References. Hence, the argument that C. neoformans is an extracellular pathogen because it has an anti-phagocytic capsule that interferes with ingestion in vivo is difficult to reconcile with existing in vivo data.We agree with Dr Levitz's succinct description of the pathogenesis of C. neoformans infection, with the caveat that we believe significant replication occurs intracellularly at the edges of C. neoformans lesions. Clearly, this is an issue for further study. Finally, Dr Levitz wonders whether the accumulation of intracellular polysaccharide is a mechanism for survival inside cells or a consequence of such survival. Separation of cause and effect in this system could be exceedingly difficult. However, C. neoformans also sheds polysaccharide inside amoebae, a natural predator (J. Steenbergen, et al., unpublished). Hence, the most straightforward interpretation is that intracellular polysaccharide production is an active mechanism for intracellular survival that could have arisen as a result of selection pressure by phagocytic environmental predators. From the standpoint of C. neoformans, there might be little difference between a macrophage and an amoeba.As noted by Dr Levitz, there is no controversy in the statement that C. neoformans is both an intracellular and extracellular pathogen. What is uncertain is the relative contribution of the intracellular and extracellular forms to pathogenesis. It is our view that a process that destroys macrophages must play a crucially important role in pathogenesis, as the macrophage is the main effector cell against this pathogen. The major novelty of the findings lies in the observation that the capsular polysaccharide contributes to virulence by being an intracellular agressin.