Summary: Photosynthetic picoeukaryotes contribute a significant fraction of primary production in the upper ocean. Micromonas pusilla is an ecologically relevant photosynthetic picoeukaryote, abundantly and widely distributed in marine waters. Grazing by protists may control the abundance of picoeukaryotes such as M. pusilla , but the diversity of the responsible grazers is poorly understood. To identify protists consuming photosynthetic picoeukaryotes in a productive North Pacific Ocean region, we amended seawater with living ¹⁵ N, ¹³ C‐labelled M. pusilla cells in a 24‐h replicated bottle experiment. DNA stable isotope probing, combined with high‐throughput sequencing of V4 hypervariable regions from 18S rRNA gene amplicons (Tag‐SIP), identified 19 operational taxonomic units (OTUs) of microbial eukaryotes that consumed M. pusilla . These OTUs were distantly related to cultured taxa within the dinoflagellates, ciliates, stramenopiles (MAST‐1C and MAST‐3 clades) and Telonema flagellates, thus, far known only from their environmental 18S rRNA gene sequences. Our discovery of eukaryotic prey consumption by MAST cells confirms that their trophic role in marine microbial food webs includes grazing upon picoeukaryotes. Our study provides new experimental evidence directly linking the genetic identity of diverse uncultivated microbial eukaryotes to the consumption of picoeukaryotic phytoplankton in the upper ocean.

Apicomplexans are a group of microbial eukaryotes that contain some of the most well-studied parasites, including the causing agents of toxoplasmosis and malaria, and emergent diseases like cryptosporidiosis or babesiosis. Decades of research have illuminated the pathogenic mechanisms, molecular biology, and genomics of model apicomplexans, but we know little about their diversity and distribution in natural environments. In this study we analyze the distribution of apicomplexans across a range of both host-associated and free-living environments. Using publicly available small subunit (SSU) rRNA gene databases, high-throughput environmental sequencing (HTES) surveys, and our own generated HTES data, we developed an apicomplexan reference database, which includes the largest apicomplexan SSU rRNA tree available to date and encompasses comprehensive sampling of this group and their closest relatives. This tree allowed us to identify and correct incongruences in the molecular identification of apicomplexan sequences. Analyzing the diversity and distribution of apicomplexans in HTES studies with this curated reference database also showed a widespread, and quantitatively important, presence of apicomplexans across a variety of free-living environments. These data allow us to describe a remarkable molecular diversity of this group compared with our current knowledge, especially when compared with that identified from described apicomplexan species. This is most striking in marine environments, where potentially the most diverse apicomplexans apparently exist, but have not yet been formally recognized. The new database will be useful for microbial ecology and epidemiological studies, and provide valuable reference for medical and veterinary diagnosis especially in cases of emerging, zoonotic, and cryptic infections.