Spatial organization of the kelp microbiome at micron scales

Background: Elucidating the spatial structure of host-associated microbial communities is essential for understanding taxon-taxon interactions within the microbiota and between microbiota and host. Macroalgae are colonized by complex microbial communities, suggesting intimate symbioses that likely play key roles in both macroalgal and bacterial biology, yet little is known about the spatial organization of microbes associated with macroalgae. Canopy-forming kelp are ecologically significant, fixing teragrams of carbon per year in coastal kelp forest ecosystems. We characterized the micron-scale spatial organization of bacterial communities on blades of the kelp Nereocystis luetkeana using fluorescence in situ hybridization and spectral imaging with a probe set combining phylum-, class-, and genus-level probes to localize and identify > 90% of the microbial community. Results: We show that kelp blades host a dense microbial biofilm composed of disparate microbial taxa in close contact with one another. The biofilm is spatially differentiated, with clustered cells of the dominant symbiont Granulosicoccus sp. (Gammaproteobacteria) close to the kelp surface and filamentous Bacteroidetes and Alphaproteobacteria relatively more abundant near the biofilm-seawater interface. A community rich in Bacteroidetes colonized the interior of kelp tissues. Microbial cell density increased markedly along the length of the kelp blade, from sparse microbial colonization of newly produced tissues at the meristematic base of the blade to an abundant microbial biofilm on older tissues at the blade tip. Kelp from a declining population hosted fewer microbial cells compared to kelp from a stable population. Conclusions: Imaging revealed close association, at micrometer scales, of different microbial taxa with one another and with the host. This spatial organization creates the conditions necessary for metabolic exchange among microbes and between host and microbiota, such as provisioning of organic carbon to the microbiota and impacts of microbial nitrogen metabolisms on host kelp. The biofilm coating the surface of the kelp blade is well-positioned to mediate interactions between the host and surrounding organisms and to modulate the chemistry of the surrounding water column. The high density of microbial cells on kelp blades (10(5)-10(7) cells/cm(2)), combined with the immense surface area of kelp forests, indicates that biogeochemical functions of the kelp microbiome may play an important role in coastal ecosystems

Bridging the Gaps in Freshwater Silicate Cycles: Assessing Silicate Demand and Incorporation in Freshwater Phytoplankton

Silicate (Si) is a potentially limiting nutrient, shaping the community structure of phytoplankton – the base of most freshwater food webs. The silicate cycle has been well studied in marine ecosystems, but is relatively poorly characterized in freshwater ecosystems, such as the Laurentian Great Lakes. Dissolved silicate concentrations in Lake Michigan have increased over the past 3 decades, which may be related to declines in Si-requiring phytoplankton such as diatoms. However, the mechanisms behind this remain unclear, as the increase appears to be too great to be due to diatoms alone. To better understand Si dynamics in lake ecosystems, the silicate demand and use by diverse freshwater phytoplankton taxa, including known Si-requiring taxa such as diatoms and chrysophytes as well as non-siliceous species, must be examined. Our experiments aimed to characterize growth rates (measured as change in chlorophyll a fluorescence) and Si incorporation into biogenic silicate (bSi) of the diatom Cyclotella meneghiniana, the green alga Chlamydomonas reinhardtii, and the chrysophyte Chrysocapsa sp. Cells were cultured with full-Si growth media, Si-free media, and full-Si media with the addition of Germanium dioxide (+GeO2), an inhibitor of Si incorporation. The diatom C. meneghiniana showed significantly decreased growth in both Si-free and +GeO2 treatments (n=3, p\u3c0.01). Biogenic silicate analyses showed significantly higher bSi in the full Si treatment (n=3, p\u3c0.01). Significant growth decreases were also seen for +GeO2 treatments for the chrysophyte Chrysocapsa (n=3, p\u3c0.01) and the green alga C. reinhardtii (n=3, p\u3c0.05). However, there were no significant differences in bSi, with both species having much lower bSi than C. meneghiniana. Fluorescence microscopy of cells exposed to a bSi-indicating fluorochrome, PDMPO, also showed bSi incorporation present only in C. meneghiniana. Experiments to quantify bSi incorporation rates using PDMPO and fluorescence spectroscopy in C. meneghiniana are currently underway

Incorporating freshwater macrophytes into the silicate budget for Estabrook Park Pond

The macronutrients nitrogen and phosphorus are known to limit primary productivity in freshwater ecosystems, but many important phytoplankton also use silicate. I examined the effect of Si on phytoplankton communities over the summer and determined that Si availability is important in shaping phytoplankton community composition. However, much of the pond is covered by rooted and suspended aquatic macrophytes, which account for much of the pond’s primary productivity, and their role in silicate cycling is unknown. The aim of our research was to characterize the silicate dynamics in Estabrook Pond, incorporating phytoplankton as well as macrophytes and their epiphytic algae, to create an Si budget for this small ecosystem. Fluxes between pools were examined in a degradation experiment using the macrophyte Myriophyllum during fall. We collected macrophyte and water samples from Estabrook pond which we divided among six different containers and stored in the growth chamber. We monitored silicate in three pools: Si dissolved in the water (dSi), particulate Si in the suspended community (pSi), and biogenic Si in the macrophytes (bSi). Over the course of the experiment, dSi was consistently low. BSi was initially high but decreased during the experiment. As bSi decreased, pSi increased suggesting breakdown of the macrophytes or detachment of epiphytes. Although the Si budget was not perfectly balanced, these findings suggest that aquatic macrophytes may have a strong influence over Si dynamics in Estabrook Pond along with diatoms and other Si-demanding cells within the phytoplankton