DataSheet_3_Lipid Secretion by Parasitic Cells of Coccidioides Contributes to Disseminated Disease.pdf

Coccidioides is a soil-borne fungal pathogen and causative agent of a human respiratory disease (coccidioidomycosis) endemic to semi-desert regions of southwestern United States, Mexico, Central and South America. Aerosolized arthroconidia inhaled by the mammalian host first undergo conversion to large parasitic cells (spherules, 80–100 μm diameter) followed by endosporulation, a process by which the contents of spherules give rise to multiple endospores. The latter are released upon rupture of the maternal spherules and establish new foci of lung infection. A novel feature of spherule maturation prior to endosporulation is the secretion of a lipid-rich, membranous cell surface layer shed in vivo during growth of the parasitic cells and secretion into liquid culture medium during in vitro growth. Chemical analysis of the culture derived spherule outer wall (SOW) fraction showed that it is composed largely of phospholipids and is enriched with saturated fatty acids, including myristic, palmitic, elaidic, oleic, and stearic acid. NMR revealed the presence of monosaccharide- and disaccharide-linked acylglycerols and sphingolipids. The major sphingolipid components are sphingosine and ceramide. Primary neutrophils derived from healthy C57BL/6 and DBA/2 mice incubated with SOW lipids revealed a significant reduction in fungicidal activity against viable Coccidioides arthroconidia compared to incubation of neutrophils with arthroconidia alone. Host cell exposure to SOW lipids had no effect on neutrophil viability. Furthermore, C57BL/6 mice that were challenged subcutaneously with Coccidioides arthroconidia in the presence of the isolated SOW fraction developed disseminated disease, while control mice challenged with arthroconidia alone by the same route showed no dissemination of infection. We hypothesize that SOW lipids contribute to suppression of inflammatory response to Coccidioides infection. Studies are underway to characterize the immunosuppressive mechanism(s) of SOW lipids.

球孢子菌属（Coccidioides）是一类土壤源性真菌病原体，可引发人类呼吸道传染病——球孢子菌病（coccidioidomycosis），该病流行于美国西南部、墨西哥、中美洲及南美洲的半干旱区域。哺乳动物宿主吸入气溶胶化的关节孢子（arthroconidia）后，孢子首先会转化为大型寄生细胞（球囊，spherules，直径80–100 μm），随后启动内孢子形成过程：球囊内部会产生大量内孢子，最终母球囊破裂释放内孢子，进而在肺部形成新的感染病灶。在进入内孢子形成阶段前，球囊成熟过程存在一项全新生物学特征：寄生细胞在体内生长时会分泌富含脂质的膜状细胞表层并发生脱落，而在体外培养条件下，该物质会被分泌至液体培养基中。对培养来源的球囊外壁（spherule outer wall, SOW）组分开展化学分析后发现，其主要成分为磷脂，且富含多种饱和脂肪酸，包括肉豆蔻酸、棕榈酸、反油酸、油酸及硬脂酸。核磁共振（NMR）检测结果显示，该组分中存在单糖、二糖连接的酰基甘油及鞘脂类物质，其中主要鞘脂成分为神经鞘氨醇（sphingosine）与神经酰胺（ceramide）。将从健康C57BL/6及DBA/2小鼠体内分离得到的原代中性粒细胞与SOW脂质共同孵育后，相较于仅将中性粒细胞与关节孢子孵育的对照组，其对活球孢子菌关节孢子的杀菌活性出现显著下降；而SOW脂质对中性粒细胞的存活率无显著影响。进一步实验表明，当以皮下注射方式用球孢子菌关节孢子感染C57BL/6小鼠时，若同时加入分离纯化的SOW组分，小鼠会出现播散性感染；而仅以关节孢子皮下接种的对照组小鼠则未发生感染播散。据此我们提出假说：SOW脂质可抑制机体针对球孢子菌感染的炎症应答。目前相关研究正在开展，以阐明SOW脂质的免疫抑制作用机制。