*Article* **"***Candidatus* **Mystax nordicus" Aggregates with Mitochondria of Its Host, the Ciliate** *Paramecium nephridiatum*

#### **Aleksandr Korotaev 1,**†**, Konstantin Benken 2 and Elena Sabaneyeva 1,\***


Received: 10 May 2020; Accepted: 16 June 2020; Published: 19 June 2020

**Abstract:** Extensive search for new endosymbiotic systems in ciliates occasionally reverts us to the endosymbiotic bacteria described in the pre-molecular biology era and, hence, lacking molecular characterization. A pool of these endosymbionts has been referred to as a hidden bacterial biodiversity from the past. Here, we provide a description of one of such endosymbionts, retrieved from the ciliate *Paramecium nephridiatum*. This curve-shaped endosymbiont (CS), which shared the host cytoplasm with recently described "*Candidatus* Megaira venefica", was found in the same host and in the same geographic location as one of the formerly reported endosymbiotic bacteria and demonstrated similar morphology. Based on morphological data obtained with DIC, TEM and AFM and molecular characterization by means of sequencing 16S rRNA gene, we propose a novel genus, "*Candidatus* Mystax", with a single species "*Ca*. Mystax nordicus". Phylogenetic analysis placed this species in Holosporales, among *Holospora*-like bacteria. Contrary to all *Holospora* species and many other *Holospora*-like bacteria, such as "*Candidatus* Gortzia", "*Candidatus* Paraholospora" or "*Candidatus* Hafkinia", "*Ca*. Mystax nordicus" was never observed inside the host nucleus. "*Ca*. Mystax nordicus" lacked infectivity and killer effect. The striking peculiarity of this endosymbiont was its ability to form aggregates with the host mitochondria, which distinguishes it from *Holospora* and *Holospora*-like bacteria inhabiting paramecia.

**Keywords:** symbiosis; *Paramecium*; ciliates; *Holospora*-like bacteria; host–parasite interactions; 16S rRNA gene; full-cycle rRNA approach; TEM; fluorescence in situ hybridization
