**Simultaneous Genome Sequencing of** *Prosthecochloris ethylica* **and** *Desulfuromonas acetoxidans* **within a Syntrophic Mixture Reveals Unique Pili and Protein Interactions**

#### **John A. Kyndt 1,\*, Jozef J. Van Beeumen 2 and Terry E. Meyer 3,**†


Received: 1 November 2020; Accepted: 4 December 2020; Published: 7 December 2020

**Abstract:** Strains of *Chloropseudomonas ethylica*, 2-K, N2, and N3 are known to be composed of a syntrophic mixture of a green sulfur bacterium and a sulfur-reducing colorless component. Upon sequence analysis, the green sulfur photosynthetic bacterial component of strain N3 was dominant and was readily sequenced, but the less abundant sulfur-reducing bacterial component was apparent only when analyzed by metagenomic binning. Whole-genome comparison showed that the green bacterium belonged to the genus *Prosthecochloris* and apparently was a species for which there was no genome sequence on file. For comparison, we also sequenced the genome of *Prosthecochloris* sp. DSM 1685, which had previously been isolated from the 2-K mixture in pure culture and have shown that all three *Prosthecochloris* genomes belong to a new species, which we propose to be named *Prosthecochloris ethylica* comb. nov. Whole genomes were also sequenced for the isolated *Desulfuromonas* strains DSM 1675 (from strain 2-K) and DSM 1676 (from strain N2) and shown to be nearly identical to the genome found in the N3 mixture. The genome of the green sulfur bacterium contains large genes for agglutination proteins, similar to the ones proposed to be involved in larger photosynthetic consortia of *Chlorochromatium aggregatum*. In addition, we also identified several unique "tight adhesion (tad)" pili genes that are presumably involved in the formation of cell–cell interactions. The colorless component, on the other hand, contained a unique large multiheme cytochrome C and unique genes for e-pili (geopilin) formation, genetically clustered with a conserved ferredoxin gene, which are all expected to play an electron transfer role in the closed sulfur cycle in the syntrophic mixture. The findings from the simultaneous genome sequencing of the components of *Cp. ethylica* have implications for the phenomenon of direct interspecies interactions and coupled electron transfer in photosynthetic symbionts. The mechanisms for such interactions appear to be more common in the environment than originally anticipated.

**Keywords:** green sulfur bacteria; syntrophy; e-pili; adhesion protein; photosynthetic symbionts; large multiheme cytochrome; metagenomic binning
