Neutrophil-mediated antifungal activity against highly virulent Cryptococcus gattii strain R265

Abstract

Vaccine-induced immune responses, including neutrophil, macrophage, and T-cell responses, ameliorate cryptococcosis caused by Cryptococcus gattii. However, whether neutrophils can exert fungicidal activity against C. gattii remains to be elucidated. Therefore, in this study, we investigated the neutrophil-mediated fungicidal effect against C. gattii R265 in vitro and compared it to the related fungal pathogen, Cryptococcus neoformans standard strain H99. We found that neutrophils recognized, phagocytosed, and killed C. gattii R265 in the presence of fresh mouse serum. This antifungal effect required phagocytosis and serine protease activity but not nicotinamide adenine dinucleotide phosphate oxidase activity. We also demonstrated that C. gattii R265 was more resistant to oxidative and nitrosative stress than C. neoformans H99. Together, these findings indicate that neutrophils can exert fungicidal activity against highly virulent C. gattii, at least under in vitro conditions.

Contribution of Laccase Expression to Immune Response against Cryptococcus gattii Infection

“…We evaluated the percentage of phagocytosis of lac1⌬, lac1⌬::LAC1, and parental C. gattii R265 strains stained with calcofluor white by neutrophils by use of flow cytometry analysis. As described in a related study (24), we found that C. gattii R265 could be engulfed by neutrophils ( Fig. 8A and B).…”

“…Laccase expression in C. gattii is required to inhibit neutrophil-mediated antifungal activity in vitro. One recent study indicated that neutrophils can mediate antifungal activity against the highly virulent C. gattii strain R265 in vitro (24). Because the laccase enzyme contributes to the inhibition of neutrophil recruitment in the lungs at the early stage of C. gattii infection, we further investigated whether the expression of laccase in C. gattii might inhibit phagocytosis and neutrophil fungicidal activity in vitro.…”

“…The possible mechanisms by which laccase inhibits antifungal activities might be due to the suppression of respiratory burst or oxidative stress in phagocytes, as previously described in a model of C. neoformans (14,35). It has been suggested that C. gattii is more resistant to oxidative stress than C. neoformans and that oxidative killing is not effective for sterilization of C. gattii R265 (24). It is likely that different mechanisms may be required for the neutrophil-mediated antifungal effects on C. gattii and C. neoformans.…”

“…Moreover, we observed that the percentage of phagocytosis and killing by neutrophils in vitro was much more pronounced in the laccase-deficient strain, indicating that laccase expression allows C. gattii R265 to resist neutrophilmediated killing to some extent. Although a recent study indicated that neutrophils can recognize, phagocytose, and kill a highly virulent C. gattii R265 strain (24), it has been suggested that the killing of C. gattii R265 by neutrophils may not be efficient for fungal sterilization. Likewise, related research suggested that IL-17 enhances the host defense against pulmonary infection with C. neoformans by affecting inflammatory cell recruitment but without affecting mouse survival (34).…”

Vaccines and Protective Immune Memory against Cryptococcosis

“…34) CPs enable cryptococcal cells to evade anticryptococcal immune systems. 35,36) We demonstrated that capsular cryptococcal cells could not be recognized by immune cells without opsonization and that acapsular mutants are immediately engulfed by innate immune cells. 14,37,38) Cryptococcal cells can be opsonized with anti-GXM antibody, and opsonized capsular cells are phagocytized by innate immune cells.…”