Immunization of mice by intracutaneous inoculation with viable virulent Cryptococcus neoformans: immunological and histopathological parameters

Abstract

Immune responses, including protection and delayed hypersensitivity, were evaluated in experimental murine cryptococcosis. Mice were immunized by the intracutaneous inoculation of viable virulent Cryptococcus neoformans yeasts. Response to the cutaneous infection was evaluated histologically and by cultural assays of the internal organs, as well as by intravenous challenge with the same strain. Protection was assessed by survival, histopathology, and quantitative organ culture. The intracutaneous inoculation of cryptococci resulted in a local inflammatory response that effectively limited dissemination of the organisms systemically and induced the development of delayed hypersensitivity demonstrable with a membrane extract of C. neoformans and with soluble cytoplasmic substances. A protective response was induced by the cutaneous inoculation of cryptococci as well, in that immunized animals survived longer, with about 25% of the challenged group ridding themselves completely of the cryptococci. Protection could be demonstrated by cultural analyses, but all animals, whether control or immunized, allowed considerable multiplication of the inoculum during the first 4 weeks after intravenous challenge. It would appear, therefore, that the protective mechanism(s) required additional antigenic stimulation before it could eventually function to eliminate all cryptococci from tissues. Histologically, there were no differences in pathology of the internal organs between immunized and unimmunized animals. Although the model described herein for the induction of immune responses in murine cryptococcosis has at least one drawback, viz., the presence of cryptococci in the skin lesion of many animals throughout the duration of the experiment, it does have the advantage that the immune responses were stimulated by a virulent strain and only minimal dissemination occurred. Therefore, lymphocytes could be removed from animals that were not contaminated with cryptococci for in vitro and in vivo transfer.

Disseminated cryptococcosis including osteomyelitis in a horse

“…There has been very little evaluation of how different routes of cryptococcal inoculation cause infection, and strain and host factors are likely to significantly affect the pattern of dissemination. One study suggests that intracutaneous inoculation is less likely to result in systemic infection, 22 but more work is needed to clarify this. The authors hypothesise that both bone lesions occurred secondary to haematogenous spread from the respiratory tract, rather than a penetrating wound leading to disseminated infection.…”

Decreased Resistance to Primary IntravenousCryptococcus neoformansInfection in Aged Mice despite Adequate Resistance to Intravenous Rechallenge

“…Inbred young adult mice infected systemically with a lethal inoculum of C. neoformans exhibit a profound meningoencephalitis. Typically, brain yeast burdens are Ͼ10 8 , and histological examination of brain tissue shows severe necrosis and edema (16,39,43). Moribund C. neoformans-infected mice have marked loss of appetite and weight and exhibit hydrocephaly.…”

L3T4(CD4)-, Lyt-2(CD8)- and Mac-1(CD11b)-phenotypic leukocytes in murine cryptococcal meningoencephalitis

“…The histopathology of murine cryptococcosis is well studied with conventional methods [5,10]. Immunohistological differentiation of leucocyte subsets has been performed for the lung [11,12,13] after intratracheal infection with C. neoformans and in human pulmonary cryptococcosis [14], but not for cryptococcal meningoencephalitis after intravenous infection.…”

“…The infiltration of CD4+ and CD8+ cells into the liver and brain of infected mice provide additional support to these findings. Similarly, depletion experiments showed that CD8+ cells are essential for the generation of delayed-type hypersensitivity (DTH) in murine cryptococcosis [ ci was found to be associated with better protection against secondary infection with the fungus, although no histological differences in pathology of the internal organs between immunized and unimmunized mice could be demonstrated [10]. These works, with the exception of a recent publication by Hill and Aguirre [37], focused on the importance of local immunity in the lungs against dissemination of the fungus to extrapulmonary sites.…”