目的 通过小型吸虫虫卵形态学检测、虫卵内转录间隔区2（ITS-2）序列扩增测序和流行病学调查分析，诊断并区分麝猫后睾吸虫和华支睾吸虫感染。 方法 收集经改良加藤厚涂片法（Kato-Katz法）检出的6例吸虫虫卵阳性病例的流行病学资料，连续3 d采集阳性病例粪样，采用水洗沉淀法收集虫卵进行镜检。提取虫卵DNA，采用rDNA ITS-2区域的通用引物进行PCR扩增并测序，产物序列在NCBI数据库中进行比对以鉴定虫种，采用邻接法构建基于ITS-2序列的系统进化树。根据虫卵形态学、基因序列比对结果和流行病学调查结果进行分析诊断。 结果 镜下可见，病例1~3粪样中的虫卵在外壳、形态、颜色、卵盖、肩峰、毛蚴和虫卵底部小疣等均与病例4~6的虫卵极其相似。病例1~3粪样中的虫卵长、宽、长宽比分别为（26.94 ± 2.28）μm、（14.43 ± 1.22）μm、1.88 ± 0.18，与病例4~6的（29.70 ± 1.21）μm、（14.36 ± 0.70）μm、2.07 ± 0.14比较，长和长宽比差异有统计学意义（t = 4.318、4.816，P < 0.01）；前者较后者短而宽，两种虫卵的浮德-梅勒尼指数（FMI）分别为5 676.69 ± 1 317.97和6 134.25 ± 626.27（t = 0.428，P > 0.05）。PCR和测序结果显示，病例1~3粪样中虫卵分别扩增出长度为355、362、359 bp的ITS-2片段，与麝猫后睾吸虫（GenBank登录号为MK886663）序列一致性分别为99.14%、98.86%、99.39%；病例4~6粪样中虫卵分别扩增出长度为394、395、396 bp的ITS-2片段，与华支睾吸虫（GenBank登录号为OK103575.1）序列一致性分别为100%、99.49%、100%。构建的系统进化树结果显示，病例1~3的ITS-2序列与麝猫后睾吸虫聚在一个分支上，病例4~6的ITS-2序列与华支睾吸虫聚在一个分支上。流行病学调查结果显示，病例1~3均为来自柬埔寨的外籍移民妇女，从小有食用腌制河鱼的习惯；病例4~6均为本地居民。结合虫卵形态学、分子生物学和流行病学调查结果，确定病例1~3为输入性麝猫后睾吸虫感染，病例4~6为华支睾吸虫感染。 结论 在形态学检测、流行病学调查分析的基础上，通过虫卵ITS-2序列扩增测序诊断能够有效鉴别麝猫后睾吸虫和华支睾吸虫感染。

Objective To explore the diagnosis and differentiation between Opisthorchis viverrin and Opisthorchis sinensis infection through morphological examination of small trematode eggs, egg internal transcribed space 2 (ITS-2) sequence amplification and epidemiological investigation. Methods The epidemiological data of 6 trematode egg positive cases diagnosed by the modified Kato-Katz thick smear method were collected, and positive fecal samples were collected for 3 consecutive days for collection of eggs by washing with water and sedimentation prior to microscopy. The eggs DNA was extracted, amplified and sequenced using universal primers for the rDNA ITS-2 region. The product sequences were aligned in the NCBI database to identify the species, and a phylogenetic tree based on ITS-2 sequences was constructed using the neighbor-joining method. The diagnosis was reached based on data analysis of the egg morphology, gene sequence alignment and epidemiological findings. Results Microscopically, the eggs in the fecal samples of cases 1 to 3 were similar to those of cases 4 to 6 in the shell, morphology, color, egg cover, acromion, hair and small wart at the bottom of the eggs. The egg length, width and aspect ratio in the fecal samples from cases 1 to 3 were (26.94 ± 2.28) μm, (14.43 ± 1.22) μm, 1.88 ± 0.18. Compared with (29.70 ± 1.21) μm, (14.36 ± 0.70) μm and 2.07 ± 0.14 in cases 4 to 6. The difference between length and aspect ratio was significant (t = 4.318, 4.816; P < 0.01). The former is shorter and wider than the latter, The Forde-Meleni index (FMI) of the two eggs was 5 676.69 ± 1 317.97 and 6 134.25 ± 626.27 (t = 0.428, P > 0.05). In cases 1 to 3, PCR and sequencing results showed that the ITS-2 fragments were 355, 362 and 359 bp, respectively, with the sequence identity of 99.14%, 98.86% and 99.39% to C. sinensis (GenBank No. MK886663). In cases 4 to 6, ITS-2 gene fragments were 394, 395 and 396 bp, respectively, with the sequence identity of 100%, 99.49% and 100% to O. viverrin (GenBank No. OK103575.1). The phylogenetic tree showed that the ITS-2 sequence of the eggs in cases 1 to 3 clustered on a branch with C. muscidiode and the ITS-2 sequence of the eggs in cases 4 to 6 clustered on a branch with C. sinensis. The epidemiological investigation results showed that cases 1 to 3 were all foreign migrant women from Cambodia who had the habit of eating pickled freshwater fish since childhood; cases 4 to 6 were all local farmers. Combining the morphological, molecular biology and epidemiological findings, cases 1 to 3 were diagnosed as imported testicular fluke infection, and cases 4 to 6 were C. sinensis infection. Conclusion Combining morphological detection as well as epidemiological investigation and analysis, trematode egg ITS-2 amplification and sequencing could effectively differentiate C. sinensis and O. viverrin in infection.