Anthropogenic carbon dioxide (CO2) emissions drive climate change and pose one of the major challenges of our century. The effects of increased CO2 in the form of ocean acidification (OA) on the communities of marine planktonic eukaryotes in tropical regions such as the Timor Sea are barely understood. Here, we show the effects of high CO2 (mean ± SD pCO2 = 1823 ± 161 μatm and pHT = 7.46 ± 0.05) versus in situ CO2 (504 ± 42 µatm, 7.95 ± 0.04) seawater on the community composition of marine planktonic eukaryotes after 3 and 48 h of treatment exposure in a shipboard microcosm experiment. Illumina sequencing of the V9 hypervariable region of 18S rRNA (gene) was used to study the eukaryotic community composition. Increased CO2 significantly suppressed the relative abundances of different eukaryotic operational taxonomic units (OTUs), including important primary producers, although the chlorophyll a concentration remained constant. OA effects on eukaryotes were consistent between total (DNA-based) and active (cDNA-based) taxa after 48 h, e.g. for the diatoms Trieres chinensis and Stephanopyxis turris. Effects of OA on the relative abundances of OTUs were often species- or even ecotype-specific, and the incubation selectively allowed for detection of the OA-sensitive OTUs that benefitted the most from incubation in a closed bottle, as containment effects on the community structure were evident after 48 h. Many OTUs were adversely affected by sudden decreases of seawater pH, suggesting high sensitivity to OA at the base of the tropical marine biodiversity and difficult-to-predict outcomes for food-web functioning in the future ocean.