Metabarcoding reveals unique microbial mat communities and evidence of biogeographic influence in low-oxygen, high-sulfur sinkholes and springs.

High-sulfur, low-oxygen environments formed by underwater sinkholes and springs create unique habitats populated by microbial mat communities. To explore the diversity and biogeography of these mats, samples were collected from three sites in Alpena, Michigan, one site in Monroe, Michigan, and one site in Palm Coast, Florida. Our study investigated previously undescribed eukaryotic diversity in these habitats and further explored their bacterial communities. Mat samples and water parameters were collected from sulfur spring sites during the spring, summer, and fall of 2022. Cyanobacteria and diatoms were cultured from mat subsamples to create a culture-based DNA reference library. Remaining mat samples were used for metabarcoding of the 16S and rbcL regions to explore bacterial and diatom diversity, respectively. Analyses of water chemistry, alpha diversity, and beta diversity articulated a range of high-sulfur, low-oxygen habitats, each with distinct microbial communities. Conductivity, pH, dissolved oxygen, temperature, sulfate, and chloride had significant influences on community composition but did not describe the differences between communities well. Chloride concentration had the strongest correlation with microbial community structure. Mantel tests revealed that biogeography contributed to differences between communities as well. Our results provide novel information on microbial mat composition and present evidence that both local conditions and biogeography influence these unique communities.

Unbiased analyses of ITS folding motifs in a taxonomically confusing lineage: Anagnostidinema visiae sp. nov. (cyanobacteria).

Cyanobacteria are diverse prokaryotic, photosynthetic organisms present in nearly every known ecosystem. Recent investigations around the world have recovered vast amounts of novel biodiversity in seldom sampled habitats. One phylogenetically significant character, the secondary folding structures of the 16S-23S ITS rDNA region, has allowed an unprecedented capacity to erect new species. However, two questions arise: Is this feature as informative as is proposed, and how do we best employ these features? Submerged sinkholes with oxygen-poor, sulfur-rich ground water in Lake Huron (USA) contain microbial mats dominated by both oxygenic and anoxygenic cyanobacteria. We sought to document some of this unique cyanobacterial diversity. Using culture-based investigations, we recovered 45 strains, of which 23 were analyzed employing 16S-23S rDNA sequences, ITS folding patterns, ecology, and morphology. With scant morphological discontinuities and nebulous 16S rDNA gene sequence divergence, ITS folding patterns were effective at articulating cryptic biodiversity. However, we would have missed these features had we not folded all the available motifs from the strains, including those with highly similar 16S rDNA gene sequences. If we had relied solely on morphological or 16S rDNA gene data, then we might well have missed the diversity of Anagnostidinema. Thus, in order to avoid conformation basis, which is potentially common when employing ITS structures, we advocate clustering strains based on ITS rDNA region patterns independently and comparing them back to 16S rDNA gene phylogenies. Using a total evidence approach, we erected a new taxon according to the International Code of Nomenclature for Algae, Fungi, and Plants: Anagnostidinema visiae.