While macroalgal microbiomes are the focus of many recent studies, there is little information about microbial spatial diversity across the thallus. Reliance on field material makes it difficult to discern whether recovered microbiomes belong to the host or its epiphytes, and technical comparisons of macroalgal samples for microbial studies are needed. Here, we use a common garden approach that avoids the problem of epiphytes, particularly at holdfasts, to examine the microbiome of Porphyra umbilicalis (strain Pum1). We used the V6 hypervariable region of the 16S rDNA with Illumina HiSeq sequencing and developed PNA clamps to block recovery of organelle V6 sequences. The common garden approach allowed us to determine differences in the microbiome at the holdfast versus blade margin. We found a notable increase in the relative abundance of Planctomycetes and Alphaproteobacteria at the holdfast, particularly of the possible symbiont Sulfitobacter sp. Nonadjacent 1.5 cm samples of blade margin had microbiomes that were not statistically different. The most abundant phylum in the overall microbiome was Proteobacteria, followed by Bacteroidetes. Because phycologists often work in remote sites, we compared three stabilization and preparation techniques and found silica gel desiccation/bead-beating and flash-freezing/lyophilization/bead-beating to be interchangeable. Core taxa (≥0.1% of sequences) across treatments were similar and accounted for ≥95% of all sequences. Finally, statistical conclusions for all comparisons were the same, regardless of which microbial community analysis tool was used: mothur or minimum entropy decomposition.

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Holopelagic Sargassum has been causing massive strandings on tropical Atlantic Ocean shorelines. After stranding, the algal biomass starts to decompose, releasing nutrients, toxic gases, and potentially introduces exogenous macro and microorganisms. Describing the microbiome associated with Sargassum, and how it changes after stranding is important in identifying potential microbial introductions to coastal environments, as well as sources of potential biotechnological resources. In this study, stranding simulation experiments were done for S. fluitans III and S. natans VIII on shipboard. Samples for microbiome identification were taken at 0 hr, just after removing healthy Sargassum from the seawater, and after 24 and 48 hrs of stranding simulation under environmental conditions. The bacterial community was identified through sequencing of 16S rRNA gene V3-V4 hypervariable regions, generating a total of 2,005 Amplicon Sequence Variants (ASVs). Of those, 628 were shared between Sargassum species. The stranding simulation changed the microbial community and only 30, out of 2,005 ASVs, persisted throughout the experiment. Phototrophs were in the main functional group at 0 hr, shifting to chemoheterotrophs within the first 24 hrs of exposure of Sargassum to air conditions. The most abundant orders Microtrichales and Rhodobacterales at 0 hr, were replaced after 24 hrs of exposure by Alteromonadales and Vibrionales, the latter representing up to 91% of the relative abundance in the bacterial community. These findings suggest that after stranding, the Sargassum microbiome goes through dysbiosis, and its biomass could become a fertile ground for potentially pathogenic bacteria.

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