Metagenomic Insights Reveal Unrecognized Diversity of Entotheonella in Japanese Theonella Sponges

Numerous biologically active natural products have been discovered from marine sponges, particularly from Theonella swinhoei, which is known to be a prolific source of natural products such as polyketides and peptides. Recent studies have revealed that many of these natural products are biosynthesized by Candidatus Entotheonella phylotypes, which are uncultivated symbionts within T. swinhoei. Consequently, Entotheonella is considered an untapped biochemical resource. In this study, we conducted metagenomic analyses to assess the diversity of Entotheonella in two T. swinhoei Y and two T. swinhoei W (Y and W referring to the yellow and white interior of the sponge, respectively), after separating filamentous bacteria using density gradient centrifugation. We obtained five Entotheonella metagenome-assembled genomes (MAGs) from filamentous bacteria-enriched fractions. Notably, one of these MAGs is significantly different from previously reported Entotheonella variants. Additionally, we identified closely related Entotheonella members present across different chemotypes of T. swinhoei. Thus, our metagenomic insights reveal that the diversity of Entotheonella within Theonella sponges is greater than previously recognized.