Cryptococcus neoformans Chitin Synthase 3 (Chs3) Plays a Critical Role in Dampening Host Inflammatory Responses

Abstract

Cryptococcus neoformans infections are significant causes of morbidity and mortality among AIDS patients and the third most common invasive fungal infection in organ transplant recipients. One of the main interfaces between the fungus and the host is the fungal cell wall. The cryptococcal cell wall is unusual among human pathogenic fungi in that the chitin is predominantly deacetylated to chitosan. Chitosan deficient strains of C. neoformans were found to be avirulent and rapidly cleared from the murine lung. Moreover, infection with a chitosan deficient C. neoformans lacking three chitin deacetylases (cda1{Delta}2{Delta}3{Delta}) was found to confer protective immunity to a subsequent challenge with a virulent wild type counterpart. In addition to the chitin deacetylases, it was previously shown that chitin synthase 3 (Chs3) is also essential for chitin deacetylase mediated formation of chitosan. Mice inoculated with chs3{Delta} at a dose previously shown to induce protection with cda1{Delta}2{Delta}3{Delta} die within 36 hours after installation of the organism. Mortality was not dependent on viable fungi as mice inoculated with heat-killed preparation of chs3{Delta} died at the same rate as mice inoculated with live chs3{Delta}, suggesting the rapid onset of death was host mediated likely caused by an over exuberant immune response. Histology, cytokine profiling, and flow cytometry indicates a massive neutrophil influx in the mice inoculated with chs3{Delta}. Mice depleted of neutrophils survived chs3{Delta} inoculation indicating that death was neutrophil mediated. Altogether, these studies lead us to conclude that Chs3, along with chitosan, plays critical roles in dampening cryptococcal induced host inflammatory responses.\n\nIMPORTANCECryptococcus neoformans is the most common disseminated fungal pathogen in AIDS patients, resulting in [~]200,000 deaths each year. There is a pressing need for new treatments for this infection, as current antifungal therapy is hampered by toxicity and/or the inability of the hosts immune system to aid in resolution of the disease. An ideal target for new therapies is the fungal cell wall. The cryptococcal cell wall is different than many other pathogenic fungi in that it contains chitosan. Strains that have decreased chitosan are less pathogenic and strains that are deficient in chitosan are avirulent and can induce protective responses. In this study we investigated the host responses to chs3{Delta}, a chitosan-deficient strain, and found mice inoculated with chs3{Delta} all died within 36 hours and death was associated with an aberrant hyperinflammatory immune response driven by neutrophils, indicating that chitosan is critical in modulating the immune response to Cryptococcus.