A case study of rhodolith beds associated with offshore NW Gulf of Mexico hard banks: viewing rhodoliths as diversity-rich holobionts

Rhodolith beds offshore Louisiana in the NW Gulf of Mexico are associated with salt domes, unique deep bank habitats at 55-75m depth on the continental shelf. Seven sampling expeditions following the 2010 Deepwater Horizon (DWH) oil spill to 2 rhodolith bed habitats revealed a drastic macroalgal die-off that has persisted up to this day (last collections from September 2014) in locations that were diversity-rich pre-DWH. Laboratory observations of bare, denuded, and apparently “dead” rhodoliths collected post-DWH and placed in 75-liter microcosms in our lab, have shown macroalgal regeneration within 3 weeks. Metabarcoding of endolithic rhodolith environmental DNAs with newly designed primers for tufA recovered a wide microbiotal diversity of photosynthetic organisms including numerous seaweed species comprising red, green, brown (and other Ochrophyta) algae, as well as other phototrophic lineages (e.g. Cyanobacteria). Metabarcoding of 16S V4 from the same endolithic rhodolith DNAs used for tufA metabarcoding recovered in-depth microbiotal taxon coverage, including abundant prokaryotic (phototrophic Cyanobacteria, heterotrophic Bacteria and Archaea), and eukaryotic phototrophic diversity. 16S V4 recovered a similar proportion of phototrophs within rhodoliths pre- and post-DWH suggesting that although not visible in the field, seaweeds are still present in the NWGMx in the form of ‘resting’ microscopic stages (dormant spores and filaments) within the CaCO₃ of rhodoliths. Using tufA and 16S metabarcoding we were able to link the taxonomic identity of the “invisible,” cryptic (hidden) parts of a macroalga that are part of a rhodolith’s CaCO₃ microbiota, with their corresponding “visible” macroscopic thalli through Sanger sequencing of plastid tufA and plastid 16S. We are continuing to characterize rhodoliths as holobionts critical for the cycling of macroalgal communities and as seedbank reservoirs and refugia of dormant microscopic stages of macroalgae. This research also illustrates and illuminates newly-found benthic life history stages of ecologically important primary producers that also cause harmful algal blooms (Dinoflagellata, Haptophyta), such as the formation of red tides, fish kills, or shellfish poisoning events in the Gulf of Mexico.