Cyanobacteria and extracellular polymeric substances (EPS) in peritidal pustular microbial mats have a two-billion-year-old fossil record. To understand the composition, production, degradation, and potential role of EPS in modern analogous communities, we sampled pustular mats from Shark Bay, Australia and analyzed their EPS matrix. Biochemical and microscopic analyses identified sulfated organic compounds as major components of mat EPS. Sulfur was more abundant in the unmineralized regions with cyanobacteria and less prevalent in areas that contained fewer cyanobacteria and more carbonate precipitates. Sequencing and assembly of the pustular mat sample resulted in 83 high-quality metagenome-assembled genomes (MAGs). Metagenomic analyses confirmed cyanobacteria as the primary sources of these sulfated polysaccharides. Genes encoding for sulfatases, glycosyl hydrolases, and other enzymes with predicted roles in the degradation of sulfated polysaccharides were detected in the MAGs of numerous clades including Bacteroidetes, Chloroflexi, Hydrogenedentes, Myxococcota, Verrucomicrobia, and Planctomycetes. Measurable sulfatase activity in pustular mats and fresh cyanobacterial EPS confirmed the role of sulfatases in the degradation of sulfated EPS. These findings suggest that the synthesis, modification, and degradation of sulfated polysaccharides influence microbial interactions, carbon cycling, and biomineralization processes within peritidal pustular microbial mats.

Pustular mats from Shark Bay, Western Australia, host complex microbial communities bound within an organic matrix. These mats harbour many poorly characterized organisms with low relative abundances (<1%), such as candidate phyla Hydrogenedentota and Sumerlaeota. Here, we aim to constrain the metabolism and physiology of these candidate phyla by analyzing two representative metagenome‐assembled genomes (MAGs) from a pustular mat. Metabolic reconstructions of these MAGs suggest facultatively anaerobic, chemoorganotrophic lifestyles of both organisms and predict that both MAGs can metabolize a diversity of carbohydrate substrates. Ca. Sumerlaeota possesses genes involved in degrading chitin, cellulose and other polysaccharides, while Ca. Hydrogenedentota can metabolize cellulose derivatives in addition to glycerol, fatty acids and phosphonates. Both Ca. phyla can respond to nitrosative stress and participate in nitrogen metabolism. Metabolic comparisons of MAGs from Shark Bay and those from various polyextreme environments (i.e., hot springs, hydrothermal vents, subsurface waters, anaerobic digesters, etc.) reveal similar metabolic capabilities and adaptations to hypersalinity, oxidative stress, antibiotics, UV radiation, nitrosative stress, heavy metal toxicity and life in surface‐attached communities. These adaptations and capabilities may account for the widespread nature of these organisms and their contributions to biofilm communities in a range of extreme surface and subsurface environments.

Here we review the application of molecular biological approaches to mineral precipitation in modern marine microbialites. The review focuses on the nearly two decades of nucleotide sequencing studies of the microbialites of Shark Bay, Australia; and The Bahamas. Molecular methods have successfully characterized the overall community composition of mats, pinpointed microbes involved in key metabolisms, and revealed patterns in the distributions of microbial groups and functional genes. Molecular tools have become widely accessible, and we can now aim to establish firmer links between microbes and mineralization. Two promising future directions include “zooming in” to assess the roles of specific organisms, microbial groups, and surfaces in carbonate biomineralization and “zooming out” to consider broader spans of space and time. A middle ground between the two can include model systems that contain representatives of important microbial groups, processes, and metabolisms in mats and simplify hypothesis testing. These directions will benefit from expanding reference datasets of marine microbes and enzymes and enrichments of representative microbes from mats. Such applications of molecular tools should improve our ability to interpret ancient and modern microbialites and increase the utility of these rocks as long-term recorders of microbial processes and environmental chemistry.

Pustular microbial mats in Shark Bay, Western Australia, are modern analogs of microbial systems that colonized peritidal environments before the evolution of complex life. To understand how these microbial communities evolved to grow and metabolize in the presence of various environmental stresses, the horizontal gene transfer (HGT) detection tool, MetaCHIP, was used to identify the horizontal transfer of genes related to stress response in 83 metagenome-assembled genomes from a Shark Bay pustular mat. Subsequently, maximum-likelihood phylogenies were constructed using these genes and their most closely related homologs from other environments in order to determine the likelihood of these HGT events occurring within the pustular mat. Phylogenies of several stress-related genes—including those involved in response to osmotic stress, oxidative stress and arsenic toxicity—indicate a potentially long history of HGT events and are consistent with these transfers occurring outside of modern pustular mats. The phylogeny of a particular osmoprotectant transport gene reveals relatively recent adaptations and suggests interactions between Planctomycetota and Myxococcota within these pustular mats. Overall, HGT phylogenies support a potentially broad distribution in the relative timing of the HGT events of stress-related genes and demonstrate ongoing microbial adaptations and evolution in these pustular mat communities.