目的 通过对多房棘球蚴病（AE）患者术前和术后、AE和细粒棘球蚴病（CE）患者间的外泌体微小RNA（miRNA）差异表达分析，评价其在棘球蚴病诊断中的应用价值。 方法 采集临床确诊的3例CE患者、4例AE患者的术前、术后（术后3~4周采样）和3位健康者全血样品，48例具AE典型病灶的患者、10例具CE典型病灶的患者（病灶均经B超检测）和39位B超阴性人群全血样品，离心分离血清。采用排阻和超滤方法分离血清中的外泌体进行富集，提取RNA进行测序，利用miRDeep2对miRNA进行比对和表达量分析，使用edgeR软件对患者术前和术后的外泌体miRNA进行差异表达分析。设计分子探针，qPCR筛选表达丰度较高的棘球蚴来源的外泌体miRNA，评价其在AE的诊断价值以及人来源的差异miRNA在棘球蚴病鉴别的诊断价值。人来源外泌体miRNA相对表达倍数的比较采用t检验。 结果 外泌体在透射电镜下呈大小不一、分布不均的圆形或类圆形囊泡状结构，直径为30~150 nm，具有明显的膜性结构。AE患者术前血清样品的外泌体中共发现了83个棘球蚴来源的miRNA，其中有64个为已知，19个为新鉴定序列。多数miRNA在血清中的表达量较低，其中emu-let-7-5p的表达量相对较高。选用qPCR初步验证表达丰度较高的2个miRNA（emu-miR-71-5p和emu-miR-10-5p）和已被报道具有诊断价值的emu-let-7-5p进行进一步验证，结果显示emu-let-7-5p的检出率为20.83%（10/48）；emu-miR-71-5p在阳性样品和阴性样品中均有检出，检出率分别为87.50%（42/48）、30.76%（12/39）；emu-miR-10-5p仅在52.08%（25/48）的阳性样品中检出。获得8个差异表达的人来源的外泌体miRNA，选取表达丰度较高的5个进行评价，其中3个（has-miR-183-5p，has-miR-222-3p和has-miR-196a-5p）在CE阳性血清中平均表达水平分别为355、299、213，是阴性对照的39.5倍、10.0倍、23.6倍，差异有统计学意义（t = 7.15、5.45、6.26，均P < 0.01）。 结论 棘球蚴来源的外泌体miRNA因在血清中痕量表达，作为棘球蚴病的诊断标记物仍存在局限性；人来源的3个外泌体miRNA在CE诊断中具有一定的研究价值。

Objective To compare the differential expression of exosomal microRNAs (miRNAs) among alveolar echinococcosis (AE) patients pre- and post- surgery, and between AE and cystic echinococcosis (CE) patients, so as to investigate the value of exosomal miRNAs for diagnosis of echinococcosis. Methods Whole blood samples were collected from pre-surgery and 3 to 4 weeks post-surgery of 3 patients with clinically definitively diagnosed CE and 4 patients with clinically definitively diagnosed AE, 3 healthy individuals, and from 48 patients with typical AE lesions, 10 patients with typical CE lesions (all lesions detected by B-mode ultrasound), and 39 individuals negative for B-mode ultrasound. Whole blood samples were centrifuged and serum samples were collected. Exosomes were isolation and enriched using size exclusion chromatography and ultrafiltration, and RNA was extracted from exosomes for sequencing. MiRNAs were identified using miRDeep2, and miRNAs expression was quantified. The differential expression of exosomal miRNAs from patients was compared pre- and post-surgery using the edgeR software. Molecular probes were designed, and Echinococcus-derived exosomal miRNAs with high abundance were screened with qPCR assay and evaluated for their values in diagnosis of AE. In addition, the value of humans-derived differentially expressed miRNAs in differential diagnosis of echinococcosis was assessed. The comparison of relative expression levels of miRNA in human derived extracellular vesicles was performed using t-test. Results Exosomes appeared circular or circular-like vesicular structures of varying sizes and uneven distribution under a transmission electron microscope, with diameters ranging from 30 to 150 nm and distinct membranous structures. A total of 83 Echinococcus-derived miRNAs were identified in the exosomes from AE patients’ pre-surgical serum samples, including 64 known miRNAs and 19 novel miRNAs, and most miRNAs had low expression in serum samples, with emu-let-7-5p showing relatively high expression. Two miRNAs with high abundance (emu-miR-71-5p and emu-miR-10-5p) screened by qPCR assay and emu-let-7-5p that had been reported to exhibit a diagnostic value were included for further verification. The detection of emu-let-7-5p was 20.83% (10/48)；emu-miR-71-5p was detected in both positive and negative samples, with detection rates of 87.50% (42/48) and 30.76% (12/39), respectively; while emu-miR-10-5p was only detected in 52.08% (25/48) of positive samples. A total of 8 differentially expressed humans-derived exosomal miRNAs were identified, and 5 were selected for evaluation based on their expression abundance. Among them, 3 miRNAs (has-miR-183-5p, has-miR-222-3p and has-miR-196a-5p) had average expression levels of 355, 299, and 213 in CE positive serum samples, which were 39.5, 10.0, and 23.6 times higher than negative controls, respectively, and the difference was statistically significant (t = 7.15, 5.45, 6.26, all P < 0.01). Conclusion Due to trace levels in serum, Echinococcus-derived exosomal miRNAs still have limitations as diagnostic markers for echinococcosis. Three humans-derived exosomal miRNAs have potential in CE diagnosis.