# Zoothamnium mariella sp. nov., a marine, colonial ciliate with an atypcial growth pattern, and its ectosymbiont Candidatus Fusimicrobium zoothamnicola gen. nov., sp. nov

## TL;DR

A new marine ciliate species and its unique bacterial ectosymbiont were discovered in the Mediterranean Sea, characterized by their unusual growth and symbiotic relationship.

## Contribution

Discovery and formal description of a new ciliate species and its novel ectosymbiotic bacterium, including their morphology, genetics, and ecology.

## Key findings

- Zoothamnium mariella sp. nov. has a unique colony structure with three distinct zooid types and a specific oral ciliature pattern.
- Candidatus Fusimicrobium zoothamnicola is a new ectosymbiont genus that covers the ciliate's surface and forms a distinct phylogenetic clade.
- The ciliate-bacterium pair thrives in temperate, oxic marine environments and can be cultivated in flow-through systems.

## Abstract

Ciliates are unicellular eukaryotes, regularly involved in symbiotic associations. Symbionts may colonize the inside of their cells as well as their surface as ectosymbionts. Here, we report on a new ciliate species, designated as Zoothamnium mariella sp. nov. (Peritrichia, Sessilida), discovered in the northern Adriatic Sea (Mediterranean Sea) in 2021. We found this ciliate species to be monospecifically associated with a new genus of ectosymbiotic bacteria, here proposed as Candidatus Fusimicrobium zoothamnicola gen. nov., sp. nov. To formally describe the new ciliate species, we investigated its morphology and sequenced its 18S rRNA gene. To demonstrate its association with a single species of bacterial ectosymbiont, we performed 16S rRNA gene sequencing, fluorescence in situ hybridization, and scanning electron microscopy. Additionally, we explored the two partners’ cultivation requirements and ecology. Z. mariella sp. nov. was characterized by a colony length of up to 1 mm. A consistent number of either seven or eight long branches alternated on the stalk in close distance to each other. The colony developed three different types of zooids: microzooids (“trophic stage”), macrozooids (“telotroch stage”), and terminal zooids (“dividing stage”). Viewed from inside the cell, the microzooids’ oral ciliature ran in 1 ¼ turns in a clockwise direction around the peristomial disc before entering the infundibulum, where it performed another ¾ turn. Phylogenetic analyses assigned Z. mariella sp. nov. to clade II of the family Zoothamnidae. The ectosymbiont formed a monophyletic clade within the Gammaproteobacteria along with two other ectosymbionts of peritrichous ciliates and a free-living vent bacterium. It colonized the entire surface of its ciliate host, except for the most basal stalk of large colonies, and exhibited a single, spindle-shaped morphotype. Furthermore, the two partners together appear to be generalists of temperate, oxic, marine shallow-water environments and were collectively cultivable in steady flow-through systems.