Several methodological issues currently hamper the study of entire trematode communities within populations of their intermediate snail hosts. Here we develop a new workflow using high-throughput amplicon sequencing to simultaneously genotype snail hosts and their infecting trematode parasites. We designed primers to amplify four snail and five trematode markers in a single multiplex PCR. While also applicable to other genera, we focused on medically and economically important snail genera within the superorder Hygrophila and targeted a broad taxonomic range of parasites within the class Trematoda. We tested the workflow using 417 Biomphalaria glabrata specimens experimentally infected with Schistosoma rodhaini, two strains of Schistosoma mansoni and combinations thereof. We evaluated the reliability of infection diagnostics, the robustness of the workflow, its specificity related to host and parasite identification, and the sensitivity to detect co-infections, immature infections and changes of parasite biomass during the infection process. Finally, we investigated its applicability in wild-caught snails of other genera naturally infected with a diverse range of trematodes. After stringent quality control the workflow allows the identification of snails to species level, and of trematodes to taxonomic levels ranging from family to strain. It is sensitive to detect immature infections and changes in parasite biomass described in previous experimental studies. Co-infections were successfully identified, opening the possibility to examine parasite–parasite interactions such as interspecific competition. Together, these results demonstrate that our workflow provides a powerful tool to analyse the processes shaping trematode communities within natural snail populations.

中文翻译：

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使用高通量扩增子测序同时对蜗牛和感染吸虫寄生虫进行基因分型

目前有几个方法问题阻碍了对其中间蜗牛宿主种群内的整个吸虫群落的研究。在这里，我们开发了一种新的工作流程，使用高通量扩增子测序同时对蜗牛宿主及其感染吸虫寄生虫进行基因分型。我们设计了引物以在单次多重 PCR 中扩增四个蜗牛和五个吸虫标记。虽然也适用于其他属，但我们专注于超纲 Hygrophila 中医学和经济上重要的蜗牛属，并针对吸虫类中广泛的寄生虫分类范围。我们使用 417个被实验性感染罗氏血吸虫（曼氏血吸虫的两种菌株）的 Biomphalaria glabrata 标本测试了工作流程及其组合。我们评估了感染诊断的可靠性、工作流程的稳健性、其与宿主和寄生虫鉴定相关的特异性，以及在感染过程中检测共感染、未成熟感染和寄生虫生物量变化的敏感性。最后，我们研究了它在自然感染各种吸虫的其他属的野生蜗牛中的适用性。经过严格的质量控制后，工作流程允许将蜗牛鉴定到物种水平，将吸虫鉴定到从科到品系的分类水平。检测以前的实验研究中描述的未成熟感染和寄生虫生物量的变化是敏感的。成功识别出合并感染，开辟了检查寄生虫 - 寄生虫相互作用的可能性，例如种间竞争。总之，这些结果表明，我们的工作流程提供了一个强大的工具来分析自然蜗牛种群中吸虫群落的形成过程。