Isolation and genomic analysis of “
Metallumcola ferriviriculae”
MK1, a Gram-positive, Fe(III)-reducing bacterium from the Soudan Underground Mine, an iron-rich Martian analog site
Authors: Hsu David , Flynn Jack R. , Schuler Christopher J. , Santelli Cara M. , Toner Brandy M. , Bond Daniel R. , Gralnick Jeffrey A. 
Published: July 2024

The Soudan Underground Mine State Park, found in the Vermilion Iron Range in northern Minnesota, provides access to a ~ 2.7 billion-year-old banded iron formation. Exploratory boreholes drilled between 1958 and 1962 on the 27th level (713 m underground) of the mine intersect calcium and iron-rich brines that have recently been subject to metagenomic analysis and microbial enrichments. Using concentrated brine samples pumped from a borehole depth of up to 55 m, a novel Gram-positive bacterium was enriched under anaerobic, acetate-oxidizing, and Fe(III) citrate-reducing conditions. The isolated bacterium, designated strain MK1, is non-motile, rod-shaped, spore-forming, anaerobic, and mesophilic, with a growth range between 24°C and 30°C. The complete circular MK1 genome was found to be 3,720,236 bp and encodes 25 putative multiheme cytochromes, including homologs to inner membrane cytochromes in the Gram-negative bacterium
Geobacter sulfurreducens
and cytoplasmic membrane and periplasmic cytochromes in the Gram-positive bacterium
Thermincola potens
. However, MK1 does not encode homologs of the peptidoglycan (CwcA) and cell surface-associated (OcwA) multiheme cytochromes proposed to be required by
T. potens
to perform extracellular electron transfer. The 16S rRNA gene sequence of MK1 indicates that its closest related isolate is
Desulfitibacter alkalitolerans
strain sk.kt5 (91% sequence identity), which places MK1 in a novel genus within the
Desulfitibacteraceae
family and
Moorellales
order. Within the
Moorellales
order, only
Calderihabitans maritimus
strain KKC1 has been reported to reduce Fe(III), and only
D. alkalitolerans
can also grow in temperatures below 40°C. Thus, MK1 represents a novel species within a novel genus, for which we propose the name “
Metallumcola ferriviriculae”
strain MK1, and provides a unique opportunity to study a cytochrome-rich, mesophilic, Gram-positive, spore-forming Fe(III)-reducing bacterium.

IMPORTANCE

The Soudan Underground Mine State Park gives access to understudied regions of the deep terrestrial subsurface that potentially predate the Great Oxidation Event. Studying organisms that have been relatively unperturbed by surface conditions for as long as 2.7 billion years may give us a window into ancient life before oxygen dominated the planet. Additionally, studying microbes from anoxic and iron-rich environments can help us better understand the requirements of life in analogous environments, such as on Mars. The isolation and characterization of “
Metallumcola ferriviriculae”
strain MK1 give us insights into a novel genus and species that is distinct both from its closest related isolates and from iron reducers characterized to date. “
M. ferriviriculae”
strain MK1 may also act as a model organism to study how the processes of sporulation and germination are affected by insoluble extracellular acceptors, as well as the impact of spores in the deep terrestrial biosphere.