我们对两种卡氏棘阿米巴菌株的细胞外囊泡(EV)进行了全面的比较分析,Neff(环境)和T4(临床)。通过透射电子显微镜进行的形态分析显示,与更多多分散的T4EV(平均=168.4nm)相比,NeffEV(平均=194.5nm)稍大。纳米粒子跟踪分析(NTA)和动态光散射验证了这些差异。电动汽车的蛋白质组学分析鉴定出1,352种蛋白质,共有1,107个,161在内夫独占,和84只在T4电动汽车。基因本体论和京都基因和基因组百科全书(KEGG)图谱揭示了不同的分子功能和生物过程,T4电动汽车富含丝氨酸蛋白酶,与其致病性一致。脂质组学分析显示,内夫电动汽车中不饱和脂质种类普遍存在,特别是三酰基甘油,磷脂酰乙醇胺(PE),和磷脂酰丝氨酸,而T4电动汽车富含二酰基甘油和二酰基甘油三甲基高丝氨酸,磷脂酰胆碱和不饱和程度较低的PE,提示脂质代谢和膜通透性的差异。代谢组学分析表明Neff电动汽车在甘油脂代谢中富集,糖酵解,和核苷酸合成,而T4电动汽车,蛋氨酸代谢。此外,EV的RNA-seq揭示了菌株之间的差异转录本,NeffEV富含与糖异生和翻译相关的转录本,提示基因调控和代谢转变,而在T4EV转录本与信号转导和蛋白激酶活性相关,表明对环境变化的快速反应。在这项小说研究中,数据整合突出了酶谱的差异,代谢过程,两种菌株中电动汽车的潜在起源揭示了A.castellanii电动汽车的多样性和复杂性,并对理解宿主-病原体相互作用和开发针对棘阿米巴相关疾病的针对性干预措施具有重要意义。IMPORTANCEA对两种具有不同毒力的棘阿米巴菌株的细胞外囊泡(EV)进行全面和全面的比较分析,aNeff(环境)和T4(临床),揭示了它们在形态和蛋白质上的惊人差异,脂质,代谢物,和成绩单水平。数据整合突出了酶谱的差异,代谢过程,以及来自这两种菌株的电动汽车的潜在不同来源,揭示了A.castellanii电动汽车的多样性和复杂性,对理解宿主-病原体相互作用有直接影响,疾病机制,并开发用于临床干预棘阿米巴相关疾病的新疗法。
We conducted a comprehensive comparative analysis of extracellular vesicles (EVs) from two Acanthamoeba castellanii strains, Neff (environmental) and
T4 (clinical). Morphological analysis via transmission electron microscopy revealed slightly larger Neff EVs (average = 194.5 nm) compared to more polydisperse
T4 EVs (average = 168.4 nm). Nanoparticle tracking analysis (NTA) and dynamic light scattering validated these differences. Proteomic analysis of the EVs identified 1,352 proteins, with 1,107 common, 161 exclusive in Neff, and 84 exclusively in
T4 EVs. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) mapping revealed distinct molecular functions and biological processes and notably, the
T4 EVs enrichment in serine proteases, aligned with its pathogenicity. Lipidomic analysis revealed a prevalence of unsaturated lipid species in Neff EVs, particularly triacylglycerols, phosphatidylethanolamines (PEs), and phosphatidylserine, while
T4 EVs were enriched in diacylglycerols and diacylglyceryl trimethylhomoserine, phosphatidylcholine and less unsaturated PEs, suggesting differences in lipid metabolism and membrane permeability. Metabolomic analysis indicated Neff EVs enrichment in glycerolipid metabolism, glycolysis, and nucleotide synthesis, while T4 EVs, methionine metabolism. Furthermore, RNA-seq of EVs revealed differential transcript between the strains, with Neff EVs enriched in transcripts related to gluconeogenesis and translation, suggesting gene regulation and metabolic shift, while in the T4 EVs transcripts were associated with signal transduction and protein kinase activity, indicating rapid responses to environmental changes. In this novel study, data integration highlighted the differences in enzyme profiles, metabolic processes, and potential origins of EVs in the two strains shedding light on the diversity and complexity of A. castellanii EVs and having implications for understanding host-pathogen interactions and developing targeted interventions for Acanthamoeba-related diseases.IMPORTANCEA comprehensive and fully comparative analysis of extracellular vesicles (EVs) from two Acanthamoeba castellanii strains of distinct virulence, a Neff (environmental) and T4 (clinical), revealed striking differences in their morphology and protein, lipid, metabolites, and transcripts levels. Data integration highlighted the differences in enzyme profiles, metabolic processes, and potential distinct origin of EVs from both strains, shedding light on the diversity and complexity of A. castellanii EVs, with direct implications for understanding host-pathogen interactions, disease mechanisms, and developing new therapies for the clinical intervention of Acanthamoeba-related diseases.