Mixotrophic protists are capable of acting both as primary producers and primary consumers at the base of the aquatic food web, thus constituting key organisms in ecosystems where they are abundant. However, their identity, abundance, ecological dynamics, and biogeochemical impact in aquatic ecosystems remain understudied in comparison to classically demarcated autotrophs or heterotrophs. In this study, we make use of fluorescently labeled prey and fluorescence‐activated cell sorting to taxonomically identify actively‐feeding individual mixotrophic flagellates from lake water. Replicated experiments were carried out to assess the performance of three different fluorescently labeled prey types and a fluorescent dye targeting food vacuoles. In the experiments, water from an oligotrophic lake was exposed independently to each type of reporter and cells were individually sorted based on fluorescent signals for predation and chlorophyll a. A total of 927 individual single cells were successfully recovered, with all fluorescent reporters exhibiting high sensitivity for putative mixotrophic taxa: overall, 87% of the occurrences could be assigned to dictyochophytes, 9% to chrysophytes, and 3% to dinoflagellates. As a result, we were able to detect cryptic diversity within pedinellid algae and report a Prorocentrum‐like freshwater occurrence. We argue that this procedure can be a valuable tool to uncover relevant and unexpected active mixotrophic species in a wider range of aquatic environments, and could easily be coupled to other techniques to describe the finer details of the trophic status of aquatic microbial communities.

中文翻译：

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荧光标记的猎物替代物与荧光激活细胞分选相结合，成功区分了湖水样本中主动摄食的混合营养生物

混合营养原生生物能够在水生食物网的基础上充当初级生产者和初级消费者，从而构成其丰富的生态系统中的关键生物。然而，与经典划分的自养生物或异养生物相比，它们的身份、丰度、生态动态和对水生生态系统的生物地球化学影响仍然没有得到充分研究。在这项研究中，我们利用荧光标记的猎物和荧光激活的细胞分选来分类识别湖水中主动进食的个体混合营养鞭毛虫。进行重复实验以评估三种不同荧光标记猎物类型和针对食物液泡的荧光染料的性能。在实验中，来自贫营养湖的水独立暴露于每种类型的报告基因，并根据捕食和叶绿素的荧光信号对细胞进行单独分类A。总共成功回收了 927 个单独的单细胞，所有荧光报告基因都对假定的混合营养类群表现出高敏感性：总体而言，87% 的事件可归属于网藻类，9% 归属于金藻类，3% 归属于甲藻类。结果，我们能够检测到足类藻类中的隐秘多样性，并报告了原甲藻属类似淡水现象。我们认为，该程序可以成为在更广泛的水生环境中发现相关和意想不到的活性混合营养物种的宝贵工具，并且可以很容易地与其他技术相结合来描述水生微生物群落营养状况的更详细信息。