Dominance of Ciliophora and Chlorophyta among phyllosphere protists of Solanaceous plants

Phyllosphere microbiota play an important role in shaping plant health and function, but most research has focused only on bacterial and fungal communities. As a result, little is known about the composition of diverse microeukaryotes in the phyllosphere, including protist predators and algae, or how they are affected by host species or co-occurring bacteria. Using universal eukaryotic primers and a PNA clamp to block amplification of the plant 18S rRNA gene, we profiled the phyllosphere and rhizosphere microbiomes of five solanaceous crop species grown in a field plot in Connecticut, USA. Phyllosphere communities of protists, like those of bacteria, were far less diverse and more variable than those of the rhizosphere. Phyllosphere samples contained hundreds of protist sequence variants from at least seven major eukaryotic lineages, of which one-fifth were not observed in bulk soil or rhizosphere samples. Phyllosphere samples were highly enriched for a few specific sequence variants representing green algae (Chlorophyta order Chlamydomonales) and ciliates (Ciliophora class Colpodea), while rhizospheres were dominated by protists from the phylum Cercozoa. Correlation analysis identified Sphingomonas spp. bacteria as central network hubs positively associated with protists in the phyllosphere and rhizosphere. These findings indicate that while leaf surfaces host a highly variable protist community of limited diversity, certain ciliates and green algae may be well-adapted for the phyllosphere habitat.