G. S. Bulmer

Control of Cryptococcus neoformans in nature by biotic factors

Two bacterial species (Pseudomonas aeruginosa and Bacillus subtilis) isolated from pigeon droppings, displayed anti Cryptococcus neoformans activity on 4 of 6 media and sterilized pigeon droppings. Acanthamoeba palestinensis trophozoites isolated from pigeon droppings ingested and killed 99.9% of C neoformans cells after 7 days of incubation. Mites and sow bugs (Metoponorthus pruinosus) isolated from pigeon droppings appear to be fungivorous. These findings suggest that many organisms that occur in pigeon droppings influence C. neoformans persistence, reproduction, morphology and distribution in nature.

In vitro effect of aqueous extract of garlic (Allium sativum) on the growth and viability of Cryptococcus neoformans.

SUMMARYThe antifungal effect of an aqueous extract of garlic was tested against 18 strains of Cryptococcus neoformans. Nine strains were clinical isolates, eight were laboratory-induced pseudohypha...

Immunization of mice with an avirulent pseudohyphal form of Cryptococcus neoformans

Mice were immunized with a viable, avirulent strain ofCryptococcus neoformans. Lymphocyte blastogenic assays showed a 10-fold increase in reactivity of sensitized spleen cells, and histopathologic examination revealed marked splenic hyperplasia. Thirty-two days after intravenous inoculation with a virulent strain ofC. neoformans, none of the control animals survived whereas 60 percent of the immunized mice were alive with no clinical evidence of disease. This animal model shows that protective immunity can be established, and once developed, provide a better model for the study of important aspects of immunity in fungal disease.

Pseudohyphal forms of Cryptococcus neoformans: Decreased survival in vivo

Three pseudohyphal isolates of Cryptococcus neoformans were inoculated intracranially into mice. Four weeks post-inoculation the animals showed no symptoms of disease and the number of viable cells per brain decreased to zero. Possible roles of pseudohyphal forms of C. neoformans in the immunology and pathogenesis of cryptococcosis are discussed.

Suppression of responses to cryptococcal antigen in murine cryptococcosis.

Subpopulations of spleen cells responsible for responsiveness and unresponsiveness to cryptococcal antigen in vitro were identified. Lymphocytes which responded in lymphocyte transformation (LT) assays were nylon wool nonadherent and theta antigen positive. These lymphocytes required the presence of an accessory cell which could be supplied by normal peritoneal exudate cells. Spleen cells taken from mice which had been infected for 3 to 15 days were tested to determine their ability to respond to cryptococcal antigen in LT assays. A minimal response was detected at the ninth day of infection. The response of infected spleen cells was attributed to a nonadherent lymphocyte. Nonadherent spleen cells of infected animals had enhanced responses after removal of adherent cells and addition of normal peritoneal exudate cells. Suppressor cells were detected in the spleens of infected mice by the 12th day of infection and thereafter. A nonadherent suppressor cell was identified, but indirect evidence suggested that an adherent cell could also be present in infected spleens.

Cryptococcus neoformans: comparisons of in vitro antifungal susceptibilities of serotypes AD and BC.

Thirty-nine isolates of Cryptococcus neoformans, nineteen serotype AD and twenty serotype BC, were assayed for susceptibility to eight antifungal agents using an in vitro agar dilution assay. Media employed were Kimmig agar and yeast nitrogen base supplemented with 10% glucose. The antifungal agents used were ketoconazole, amphotericin B, 5-fluorocytosine, nystatin, miconazole, BAY N 7133, ICI 153,066, and itraconazole. No clinically significant differences in in vitro minimum inhibitory concentrations were detected between serotypes AD and BC against any of the compounds tested. An adverse medium effect was observed in two of the assays, but the outcome of the AD/BC comparison was not affected. This is the first report in which the in vitro antifungal susceptibilities of Cryptococcus neoformans serotypes are analyzed.

A one year study on the viability of Cryptococcus neoformans in nature

Using four different assay procedures we found a 50 to 86% decrease in viability of Cryptococcus neoformans over a one year period after two towers in the Oklahoma City area were modified to exclude pigeons and weather elements, especially rain. In homogeneous pigeon dropping samples a sharp decrease in viability occurred after 7 to 9 months of storage. The decrease occurred in pigeon dropping samples maintained at the natural sites and at ambient temperature in the laboratory but not in droppings stored at −4 °C. Air sampling studies done at the natural sites before and after tower modifications also showed a decrease in C. neoformans viability. The decrease in viability cannot be explained by seasonal temperature variations that occurred at the natural sites. Our findings suggest that death of C. neoformans occurred because of lack of available intracellular water exacerbated by low humidity, exclusion of precipitation and moisture from fresh pigeon droppings.In light of our experiences, at one site in south central United States, the feasibility of controlling C. neoformans at heavily contaminated sites, other than by the use of noxious or potentially dangerous chemicals, is discussed.

Culture of Cryptococcus neoformans in the nonencapsulated state

Forty-one strains of Cryptococcus neoformans were examined after 3 days growth on a fresh and aged medium at pH 5 & pH 7 for comparison of capsule formation. Over one-half of the strains did not form visible capsules on aged medium at pH 5. Serotypes and source of isolation did not correlate with ability or inability to form capsules. Growth of C. neoformans in the nonencapsulated state makes it possible to culture many strains of C. neoformans in the form that more closely simulates the true infectious particles.