Microbiome of macroorganisms might directly or indirectly influence host development and homeostasis. Many studies focused on the diversity and distribution of prokaryotes within these assemblages, but the eukaryotic microbial compartment remains underexplored so far. To tackle this issue, we compared blocking and excluding primers to analyze microeukaryotic communities associated with Crassostrea gigas oysters. High-throughput sequencing of 18S rRNA genes variable loops revealed that excluding

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... primers performed better by not amplifying oyster DNA, whereas the blocking primer did not totally prevent host contaminations. However, blocking and excluding primers showed similar pattern of alpha and beta diversities when protist communities were sequenced using metabarcoding. Alveolata, Stramenopiles and Archaeplastida were the main protist phyla associated with oysters. In particular, Codonellopsis, Cyclotella, Gymnodinium, Polarella, Trichodina, and Woloszynskia were the dominant genera. The potential pathogen Alexandrium was also found in high abundances within some samples. Our study revealed the main protist taxa within oysters as well as the occurrence of potential oyster pathogens. These new primer sets are promising tools to better understand oyster homeostasis and disease development, such as the Pacific Oyster Mortality Syndrome (POMS) targeting juveniles.