Cyanobacterium Microcoleus in toxic benthic mats on different streambed substrates: Ecophysiology and important...

Benthic cyanobacteria, notably the genus Microcoleus, are a common contributor to benthic harmful algal blooms globally and can produce neurotoxins. Microcoleus can thrive in nutrient-limited freshwater environments, which present significant environmental and public health challenges. In May 2023, we observed Microcoleus mat growing in a small tributary of the Virgin River in Zion National Park near the Temple of Sinawava and collected benthic mats from three rock and three sandy substrate (strata) sites in the Virgin River. The overall objective of this study was to evaluate the effect of the bottom substrate (sand versus rock) on the ecophysiology of cyanobacteria, primarily Microcoleus, and other coexisting bacteria. Toxin measurements revealed that all the benthic mat samples contained anatoxin-a (ATX377.13±18.05 µg/g of wet mat) and dihydroanatoxin-a (15±0.3 µg/g of wet mat), and anatoxin-A was also present in the water column (0.377 µg/L). Low chlorophyll-a levels and microscopy results indicate that the toxins in the water flowing into the Virgin River presumably originated from benthic sources rather than from planktonic algae. Community analysis showed strong cyanobacterial dominance (>60%) in mats. Biofilms, especially those formed on sand as compared to those formed on rocks, supported greater heterotrophic bacterial diversity. A single dominant toxigenic Microcoleus genotype occurred across both strata (rock and sand) at all sampled sites, and it is closely related to the Microcoleus anatoxicus previously found in the Russian River, CA. Bottom strata type effects were most prominent in phosphorus acquisition: rock-associated heterotrophic communities showed higher expression of phosphonate utilization genes (C-P lyase) and glycerophosphodiester utilization (ugp). Samples from both substrates showed strong expression of pst/pho regulators, indicating organic phosphorus uptake. Active nitrogen fixation genes were also found in some metagenomic-assembled genomes (MAGs), suggesting internal nitrogen cycling in Microcoleus mats. Despite producing dihydroanatoxin-a, Microcoleus MAGs from this study lack the anaK gene, which is hypothesized to convert anatoxin-a to dihydroanatoxin-a. Toxic Microcoleus genomes recovered from Zion National Park encoded a complete thiamine biosynthesis pathway, including thiD. This contrasts with previous studies, which reported thiD loss in toxic Microcoleus. Overall, our results show a stable toxic Microcoleus genotype that dominates across substrates, while substrate-linked community functions between rock and sand habitats vary, especially in phosphorus acquisition.