The Complete Genome and Physiological Analysis of the Eurythermal Firmicute Exiguobacterium chiriqhucha Strain RW2 Isolated From a Freshwater Microbialite, Widely Adaptable to Broad Thermal, pH, and Salinity Ranges

Members of the genus Exiguobacterium are found in diverse environments from marine, freshwaters, permafrost to hot springs. Exiguobacterium can grow in a wide range of temperature, pH, salinity, and heavy-metal concentrations. We characterized Exiguobacterium chiriqhucha strain RW2 isolated from a permanently cold freshwater microbialite in Pavilion Lake, British Columbia using metabolic assays, genomics, comparative genomics, phylogenetics, and fatty acid composition. Strain RW2 has the most extensive growth range for temperature (4–50°C) and pH (5–11) of known Exiguobacterium isolates. Strain RW2 genome predicts pathways for wide differential thermal, cold and osmotic stress using cold and heat shock cascades (e.g., csp and dnaK), choline and betaine uptake/biosynthesis (e.g., opu and proU), antiporters (e.g., arcD and nhaC Na+/K+), membrane fatty acid unsaturation and saturation. Here, we provide the first complete genome from Exiguobacterium chiriqhucha strain RW2, which was isolated from a freshwater microbialite. Its genome consists of a single 3,019,018 bp circular chromosome encoding over 3,000 predicted proteins, with a GC% content of 52.1%, and no plasmids. In addition to growing at a wide range of temperatures and salinities, our findings indicate that RW2 is resistant to sulfisoxazole and has the genomic potential for detoxification of heavy metals (via mercuric reductases, arsenic resistance pumps, chromate transporters, and cadmium-cobalt-zinc resistance genes), which may contribute to the metabolic potential of Pavilion Lake microbialites. Strain RW2 could also contribute to microbialite formation, as it is a robust biofilm former and encodes genes involved in the deamination of amino acids to ammonia (i.e., L-asparaginase/urease), which could potentially boost carbonate precipitation by lowering the local pH and increasing alkalinity. We also used comparative genomic analysis to predict the pathway for orange pigmentation that is conserved across the entire Exiguobacterium genus, specifically, a C30 carotenoid biosynthesis pathway is predicted to yield diaponeurosporene-4-oic acid as its final product. Carotenoids have been found to protect against ultraviolet radiation by quenching reactive oxygen, releasing excessive light energy, radical scavenging, and sunscreening. Together these results provide further insight into the potential of Exiguobacterium to exploit a wide range of environmental conditions, its potential roles in ecosystems (e.g., microbialites/microbial mats), and a blueprint model for diverse metabolic processes

The Complete Genome and Physiological Analysis of the Microbialite-Dwelling Agrococcus pavilionensis sp. nov; Reveals Genetic Promiscuity and Predicted Adaptations to Environmental Stress

Members of the bacterial genus Agrococcus are globally distributed and found across environments so highly diverse that they include forests, deserts, and coal mines, as well as in potatoes and cheese. Despite how widely Agrococcus occurs, the extent of its physiology, genomes, and potential roles in the environment are poorly understood. Here we use whole-genome analysis, chemotaxonomic markers, morphology, and 16S rRNA gene phylogeny to describe a new isolate of the genus Agrococcus from freshwater microbialites in Pavilion Lake, British Columbia, Canada. We characterize this isolate as a new species Agrococcus pavilionensis strain RW1 and provide the first complete genome from a member of the genus Agrococcus. The A. pavilionensis genome consists of one chromosome (2,627,177 bp) as well as two plasmids (HC-CG1 1,427 bp, and LC-RRW783 31,795 bp). The genome reveals considerable genetic promiscuity via mobile elements, including a prophage and plasmids involved in integration, transposition, and heavy-metal stress. A. pavilionensis strain RW1 differs from other members of the Agrococcus genus by having a novel phospholipid fatty acid iso-C15:1Δ4, β-galactosidase activity and amygdalin utilization. Carotenoid biosynthesis is predicted by genomic metabolic reconstruction, which explains the characteristic yellow pigmentation of A. pavilionensis. Metabolic reconstructions of strain RW1 genome predicts a pathway for releasing ammonia via ammonification amino acids, which could increase the saturation index leading to carbonate precipitation. Our genomic analyses suggest signatures of environmental adaption to the relatively cold and oligotrophic conditions of Pavilion Lake microbialites. A. pavilionensis strain RW1 in modern microbialites has an ecological significance in Pavilion Lake microbialites, which include potential roles in heavy-metal cycling and carbonate precipitation (e.g., ammonification of amino acids and filamentation which many trap carbonate minerals)

Metagenomic analysis suggests modern freshwater microbialites harbor a core and distinct microbial community

Modern microbialites are complex microbial communities that interface with abiotic factors to form carbonate-rich organosedimentary structures whose ancestors provide the earliest evidence of life. Past studies primarily on marine microbialites have inventoried diverse taxa and metabolic pathways, but it is unclear which of these are members of the microbialite community and which are introduced from adjacent environments. Here we control for these factors by sampling the surrounding water and nearby sediment, in addition to the microbialites and use a metagenomics approach to interrogate the microbial community. Our findings suggest that the Pavilion Lake microbialite community profile, metabolic potential and pathway distributions are distinct from those in the neighboring sediments and water. Based on RefSeq classification, members of the Proteobacteria (e.g alpha and delta classes) were the dominant taxa in the microbialites, and possessed novel functional guilds associated with the metabolism of heavy metals, antibiotic resistance, primary alcohol biosynthesis and urea metabolism; the latter may help drive biomineralization. Urea metabolism within Pavilion Lake microbialites is a feature not previously associated in other microbialites. The microbialite communities were also significantly enriched for cyanobacteria and acidobacteria, which likely play an important role in biomineralization. Additional findings suggest that Pavilion Lake microbialites are under viral selection as genes associated with viral infection (e.g CRISPR-Cas, phage shock and phage excision) are abundant within the microbialite metagenomes. The morphology of Pavilion Lake microbialites changes dramatically with depth; yet, metagenomic data did not vary significantly by morphology or depth, indicating that microbialite morphology is altered by other factors, perhaps transcriptional differences or abiotic conditions. This work provides a comprehensive metagenomic perspective of the interactions and differences between microbialites and their surrounding environment, and reveals the distinct nature of these complex communities